Invitation
Welcome!
You are cordially invited to the 3rd Asian Chemical Biology Conference, which will
be held from 15 to 17 December 2014 (ACBC2014) at Ngee Ann Kongsi Auditorium,
University Town, National University of Singapore.
ACBC2014 is dedicated to promote advances in chemical biology and aims to
reflect on the significant developments of drug design, its discovery and delivery
to biological system, to discuss new ideas and trends of chemical biology, as well
as to raise the profile of chemical biology and medicinal chemistry in Asia. A wide
range of contemporary research areas in chemical biology and its applications will
be presented in this conference.
ACBC2014 serves as an important Asian forum that brings together crossdisciplinary scientists from academia, non-profit organizations, government, and
industry to communicate new research and to help translate the power of chemical
biology to advance human health. As per its tradition, the scientific committee will
ensure the high quality and diversity of the scientific programme, together with the
accessibility of the meeting to MSc and PhD students.
The best scientific researchers of ACBC2014 share the goal of using chemical
method/technology to develop a better understanding of biological processes and
to harness these processes for the common good. The objectives of the ACBC are
to promote knowledge and research in the field of biological chemistry and to
advance the relations of this discipline to other branches of science.
We look forward to welcoming you to ACBC2014 in Singapore.
Prof. Young-Tae CHANG
Chairman, the 3rd
Asian Chemical Biology Conference
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Organizing Committee
Chairman
Young-Tae Chang (NUS and SBIC, A*STAR)
Secretary
Sung-Jin Park (SBIC, A*STAR)
Nam-Young Kang (SBIC, A*STAR)
Meiling Zhang (NUS)
Committee members
Koichi Fukase (Japan)
Hisakazu Mihara (Japan)
Itaru Hamachi (Japan)
Injae Shin (Korea)
Jaehoon Yoo (Korea)
Chun-Hung Hans Lin (Taiwan)
Zhihong Guo (Hong Kong/China)
Bengang Xing (Singapore)
Anh Tuan Phan (Singapore)
Shao Q. Yao (Singapore)
James P. Tam (Singapore)
Markus Wenk (Singapore)
Ho Sup Yoon (Singapore)
Seong-Wook Yun (Singapore)
Sohail Ahmed (Singapore)
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Sponsors and Supports
Gold Sponsor
-3-
Silver Sponsor
Surpported by
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Symposium Date & Venue
Dates
December 15 (Mon) – 17 (Wed), 2014
Venue
Ngee Ann Kongsi Auditorium, Level 2
Education resource centre
University Town (U-Town), NUS
8 College Avenue West, Singapore, 138608
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General Information
Lectures
Invited Speaker lecture: 20min including discussion
Short Oral Presentation: 10min presentation (no discussion)
Invited Lectures & Short Oral Presentation
LCD projector is the standard medium for all lectures. Power point presentations are
recommended. We appreciate if you could bring your PC and connect to the switcher
during the break before your session. We ask you to preview and check your
presentations in advance. Should you bring your presentation data saved in USB
memory sticks, we will copy your file onto our computers and will check if your
presentation is displayed properly.
Poster Session
Poster presentation is scheduled from 13:00 to 14:00 on December 16 th. The
presenters should be present during the time for discussion. All posters should be
posted before 13:00 on December 15th and removed by 11:00 on December 17th.
The poster board will be prepared for each poster and materials for attaching posters
will be provided at the conference site.
Conference Language
The conference language, for both written and spoken medium will be English.
Banquet
The conference banquet will be held at 19:00 on December 15 th (Mon) at the
“Compass Ballroom” in Raffles Marina.
Shuttle Bus Time Schedule
Date
th
15 Dec
Pick-up Time
11:00 AM
11:30 AM
05:30 PM
05:35 PM
05:40 PM
09:30 PM
th
16 Dec
th
17 Dec
07:45 AM
07:50 AM
18:00 PM
18:10 PM
18:20 PM
07:45 AM
07:50 AM
01:30 PM
02:00 PM
06:00 PM
Destination
Big hotel → U-town
U-town → Raffles Marina
Destination address
U-town: Stephen Riady Center, 2 College
Avenue West, 138607
Raffles Marina: 10 Tuas West Drive Singapore
638404
Raffles Marina → Big hotel
Raffles Marina → Jurong east MRT
Big hotel → U-town
U-town → Speaker Dinner
U-town → Local dinner
U-town → Big hotel
Big hotel → U-town
U-town → City Tour
U-town → Big hotel
Big hotel → Dinner
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Dempsey
Big hotel: 200 Middle Road, Singapore 188980
Conference Program
15th December (Monday)
Time
Program
11:00 - 18:00
Registration
13:00 – 13:10
13:10 – 13:20
13:20 – 13:40
13:40 – 14:00
14:00 – 14:20
14:20 – 14:40
14:40 – 15:00
Opening Remarks
Prof. Young-Tae Chang (National University of Singapore)
Welcome Address
Prof. Patrick J. Cozzone (Singapore Bioimaging Consortium, A-STAR)
Session I. Nanobio-science and Technology for Living System
(Chairman: Prof. Chun-Hung Lin )
Deprotection chemistry-mediated protein activation in living cells
Peng Chen, Peking-Tsinghua Center for Life Sciences
Turn-ON fluorescent affinity labeling using a small bifunctional Onitrobenzoxadiazole unit
Mikiko Sodeoka, Synthetic Organic Chemistry Laboratory and CSRS, RIKEN
Detection of native proteins in live cells by iFIT (intrinsic Förster resonance
energy transfer Imaging Technology)
Sang J. Chung, Dongguk University
Nano- and Quantum-biodevices for Biomedical Applications
Yoshinobu Baba, Nagoya University
Cellular and in vivo Imaging Probes with Tunable Chemical Switches
Kazuya Kikuchi, Osaka University
15:00 – 15:20
Group Photo
15:20 – 15:50
Coffee Break
15:50 – 16:10
16:10 – 16:30
16:30 – 16:50
16:50 – 17:10
17:10 – 17:30
Session 2. DNA Structure and Biological Function
(Chairman: Prof. Koichi Fukase)
Integrative chemical biology approaches to examine histone posttranslational
modifications
Xiang David Li, The University of Hong Kong
Design and Construction of Fluorescent Nucleic Acid Systems
Byeang Hyean Kim, Pohang University of Science and Technology
Structures, Interactions, and Functions of Nucleic Acids under Molecular
Crowding Conditions
Naoki Sugimoto, Konan University
Distinct role of water in protein-protein interactions
Sihyun Ham, Sookmyung Women’s University
Metalloprotein design using genetic code expansion
Jiangyun Wang, Chinese Academy of Sciences
19:00 – 21:30
Banquet
-7-
16th December (Tuesday) AM
Time
Program
09:00 - 18:00
Registration
09:00 – 09:20
09:20 – 09:40
09:40 – 10:00
Session 3. Protein and Peptide Structure and Interaction
(Chairman: Dr. Su Seong Lee & Prof. Yan Mei Li)
Towards Pharmacological Modulation of Protein-Protein Interactions
Hyun-Suk Lim, Pohang University of Science & Technology
Peptide-based approaches for delivering exogenous molecules into cells
Shiroh Futaki, Kyoto Univeristy
Biomedical Applications of a Novel Class of High-affinity Peptides (Aptides)
Sangyong Jon, Korea Advanced Institute of Science and Technology
10:00 – 10:30
10:30 – 10:40
10:40 – 10:50
10:50 – 11:10
11:10 – 11:30
11:30 – 11:50
11:50 – 12:10
12:10 – 12:30
Coffee Break
Short Oral Presentation
(Chairman: Dr. Chai Lean Teoh)
Chai Lean Teoh, Singapore Bioimaging Consortium
Yun Kyung Kim, Korea Institute of Science and Technology (KIST)
Chemically synthetic glycopeptide vaccines
Yan-Mei Li, Tsinghua University
Robust and reliable high-throughput screening of bead-based peptide libraries
Su Seong Lee, Institute of Bioengineering and Nanotechnology
Protein
phosphorylation
of
methanoarchaeon
Methanohalophilus
portucalensis FDF1T in methanogenesis and osmoadaptation
Shih-Hsiung Wu, Academia Sinica
Development of programmable biological functionalities
Matthew W. Chang, National University of Singapore
“MicroAntibodes”: Generation of Molecular-targeting Peptides by Directed
Evolution in Conformationally Constrained Peptide libraries
Ikuo Fujii, Osaka Prefecture University
12:30 – 14:00
Lunch Break
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16th December (Tuesday) PM
13:00 – 14:00
14:00 – 14:10
14:10 – 14:20
14:20 – 14:40
14:40 – 15:00
15:00 – 15:20
15:20 – 15:40
15:40 – 16:00
Poster Session
Business Meeting (Organizing Committee)
Short Oral Presentation
(Chairman: Dr. Chai Lean Teoh)
Soonsil Hyun, Seoul National University
Satoshi Arai, WASEDA Bioscience Research Institute in Singapore (WABIOS)
Session 4. Molecular Strategies for Chemical Biology
(Chairman: Prof. Motonari Uesugi & Prof. Jong Seung Kim)
Pseudo-Natural Product Synthesis and Drug Applications
Hiroaki Suga, The University of Tokyo
Single molecule based cancer specific targeting drug delivery
Jong Seung Kim, Korea University
Chemical and Enzymatic Syntheses of Carbohydrate Oligomers
Chun-Cheng Lin, National Tsing-Hua University
How we can improve affinity of antibodies for the targets
Kouhei Tsumoto, The University of Tokyo
Intrinsic Small Molecule Fluorescent Imaging Probes towards Localized
Identification of Drug Resistant Bacterial Strains
Bengang XING, Nanyang Technological University
16:00 – 16:30
16:30 – 16:50
16:50 – 17:10
17:10 – 17:30
17:30 – 17:50
Coffee Break
Biosynthesis of Streptolidine Involved Two Unexpected Intermediates
Produced by a Dihydroxylase and a Cyclase through Unusual Mechanisms
Tsung-Lin Li, Academia Sinica
Small Molecule Tools for Cell Therapy
Motonari Uesugi, Kyoto University
Kinome profiling of 14-membered resorcylic acid lactones and identification of
drug binding sites of drugable kinases
Taebo Sim, Korea Institute of Science and Technology
A Chemical Strategy for Traceless Labeling and Immobilization of
Glycoproteins
Po-Chiao Lin, National Sun Yat-sen University
19:00 – 21:00
Local Dinner
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17th December (Wednesday)
Time
Program
09:00 - 11:00
Registration
09:00 – 09:10
09:10 – 09:20
09:20 – 09:40
09:40 – 10:00
10:00 – 10:30
10:30 – 10:50
10:50 – 11:10
11:10 – 11:30
11:30 – 11:50
Short Oral Presentation
(Chairman: Dr. Chai Lean Teoh)
Jun-Seok Lee, Korea Institute of Science and Technology (KIST)
Hyun woo Rhee, Ulsan National Institute of Science and Technology (UNIST)
Session 5. Glycan/Lipid; Biofunctional Molecules and Materials.
(Chairman: Prof. Injae Shin)
Glyco-based interplay between Helicobacter pylori and gastric epithelium
Chun-Hung Hans Lin, Academia Sinica
Glycosylation of flavonoids and polyketides using flexible glycosyltransferase
Jae Kyung Sohng, Sun Moon University
Coffee Break
Microbial glycoconjugates and lipidconjugates as immunomodulators;
Chemical synthesis and biological functions
Yukari Fujimoto, Keio University
Labeling and Visualizing Glycans with Specificity and Versatility
Xing Chen, Peking University
Bio-recognition and Bio-sensing---From big to small
Yaw-Kuen Li, National Chiao Tung Universty
Fusarisetin A, A Novel Cancer Cell Migration Inhibitor from Fusarium sp.
FN080326, and It’s Cellular Binding Protein
Jong Seog Ahn, Korea Research Institute of Bioscience and Biotechnology
11:50 – 12:00
Closing Address
12:00 – 13:20
13:30 – 17:00
19:00 – 21:00
Lunch
City Tour
Tour Dinner
- 10 -
Poster Session
13:00 – 14:00, 16th December (Wed)
Poster
No.
P-01
P-02
P-03
P-04
P-05
P-06
Title and Presenter
Engineering hemolytic peptides for intracellular delivery of biomacromolecules
Misao Akishiba, Toshihide Takeuchi, Yoshimasa Kawaguchi, Shiroh Futaki
(Kyoto University)
Plant-Based Polymeric Materials Design using a Sugar-diol as a Pendant Group
Hidenobu Takao, Yuzo Ishigaki, Keigo Aoi (Nagoya University)
Fluorescent Probes for Targeted Visualization of Temperature at Organelles in
single living cells
Satoshi Arai, Madoka Suzuki, Young-Tae Chang
(WASEDA Bioscience Research Institute in Singapore)
Dual-targeting delivery systems: selective cancer cell death and imaging
Kyung-Hwa Baek, Xizhe Tian, Injae Shin (Yonsei University)
Investigating the Mechanism of Lipid A Binding to CD14 using Molecular Dynamics
Simulations
Nils A. Berglund, Syma Khalid, Peter J. Bond ( Bioinformatics Institute, A*STAR)
Enzymatic Introduction of PEG Strands onto DNA and Effects of the PEG
Modification on Cell Adhesion
Hiroto Fujita, Kosuke Nakajima, Yuuya Kasahara, Masayasu Kuwahara
(Gunma University)
P-07
Efficient Labeling of Amino Groups by Azaelectrocyclization, Application to PET and
Fluoresecent Imaging of Biomolecules and Living Cells.
Koichi Fukase, Katsunori Tanaka (Osaka University)
P-08
Synthetic Study of Clickable Dysiherbaine
Koichi Fukushima, Masato Oikawa ( Yokohama City University)
P-09
Ligand-dependent active site closure revealed in the crystal structure of
Mycobacterium tuberculosis MenB complexed with product analogs
Haigang Song, Hoi Pang Sung, Yuk Sing Tse, Ming Jiang, Zhihong Guo
(The Hong Kong University of Science and Technology)
P-10
Study on the Target Proteins of Antitumor Macrolide Aplyronine A
Yuichiro Hirayama, Kozo Yoneda, Kota Yamagishi, Kota Tsuchiya, Takumi Chinen,
Takeo Usui, Eriko Sumiya, Motonari Uesugi, Tomohiro Suzuki, Hirokazu Kawagishi,
Masaki Kita, Hideo Kigoshi (University of Tsukuba)
P-11
Proteomic Analysis on Enhanced Tolerance of Nanoencapsulated Saccharomyces
Cerevisiae against UV-C Irradiation
Daewha Hong, Hojae Lee, Taegyun Park, Hee Chul Moon, Insung Choi
(Korea Advanced Institute of Science and Technology)
P-12
Chemoenzymatic synthesis of cyclic sialyl Lewis X 6-sulfate and analogues
Chuen-Jiuan Huang (Academia Sinica)
- 11 -
P-13
P-14
P-15
P-16
P-17
P-18
P-19
P-20
P-21
P-22
P-23
P-24
P-25
Cell-Penetrating, Dimeric a-Helical Peptides: Nanomolar Inhibitors of
HIV-1 Transcription
Sangmok Jang, Soonsil Hyun, Seoyeon Kim, Seonju Lee, Im-Soon Lee, Masanori
Baba, Yan Lee, Jaehoon Yu (Seoul National University)
Single Chirality Separation of Single-Walled Carbon Nanotube Using Flavin
Mononucleotide by Density Gradient Ultracentrifugation
Myungsu Jang, Haneul Jeong, Jinsook Sim, and Sang-Yong Ju (Yonsei University)
Protein controlled bipolar doping of a monolayer grapheme
Ji-ryang Jang, Sung Kyu Jang, Sungjoo Lee, Woo-Seok Choe
(Sungkyunkwan University)
Graphene Nanoribbons Formed by a Sonochemical Method using Flavin
Mononucleotide Assembly as a Template
Sang-Yong Ju, Woojin Yoon (Yonsei University)
Development of pancreatic β-cell probes binding to insulin for in vivo imaging
Nam Young Kang, Jungyeol Lee, Sung-Jin Park, Wut Hmone Phue, Bikram Keshari
Agrawalla, Young-Tae Chang (Singapore Bioimaging Consortium)
A New Genetically Encodable Flavinylation Tag for Screening Proteasome Inhibitors
In Living Mammalian Cells
Myeong-Gyun Kang, Hyun Woo Rhee (Ulsan National Institute of Science and
Technology)
Sending innate immune signals across the membrane: A multiscale simulation
approach to Toll-like receptor assembly
Vasileios E Kargas, Daniel A Holdbrook, Isaac Godfroy, Hang H Yin, Robert C Ford,
Peter J Bond (Bioinformatics Institute, A*STAR)
Synthesis and Binding Analysis of Thioflavin T Analogs Targeting G-Quadruplex
Yuka Kataoka, Hiroto Fujita,Yuuya Kasahara, Toshitada Yoshihara, Seiji Tobita,
Masayasu Kuwahara (Gunma University)
Identification of prion-like tau species
Dohee Kim, Sung-su Lim, Mamun Harque, Yun Kyung Kim
(Korea Institute of Science and Technology (KIST))
Peptide-mediated synthesis of palladium raspberry nanoparticles and their effect
on catalytic activities in Sonogashira cross-coupling reaction
Young-O Kim, Hyeong-Seok Jang, Yo-Han Kim, Jae Myoung You, Yoon-Sik Lee
(Seoul National University)
Efficient aerobic oxidation of alcohols to aldehydes over graphene oxide supported
ruthenium oxide catalyst under aqueous condition
Yo-Han Kim, Jung Won Kim, Yoon-Sik Lee (Seoul National University)
Crystallographic and biochemical studies of ilvC, a ketol-acid reductoisomerase,
from Streptococcus pneumoniae D39
GyuHee Kim, GyuLee Kim, Sumin Lee, Jaesook Yoon, Dong-Kwon Rhee, Sangho Lee
(Sungkyunkwan University)
Exploitation of chemistry and biology of siderophores to combat against drugresistant bacteria
Hak Joong Kim, Hwisoo Ree, Jimin Kim, Hyeon Seok Kim, Woon Young Song, Seong
Ji Choi, Mijin Sun, Jin-su An, Jae Eun Lee (Korea University)
- 12 -
P-26
Harnessing Peptides for Electrochemical Detection of Bisphenol A
Sung-Eun Kim, Jiao Yang, Misuk Cho, Ik-Keun Yoo, Youngkwan Lee, Woo-Seok Choe
(Sungkyunkwan University)
P-27
Engineering of Mono-Disperse Oligoethylene Glycols for Protein Manipulation
K. Kinbara, T. Muraoka, K. Adachi, M. Ui, S. Kawasaki, N. Sadhukhan, H. Obara, M.
Laguerre, H. Tochio, M. Shirakawa (Tohoku University)
P-28
P-29
P-30
P-31
P-32
Stylissatin A, A Cyclic Peptide That Inhibits Nitric Oxide Production from Marine
Sponge
Masaki Kita, Baro Gise, Atsushi Kawamura, Taiki Sunaba, Tito Akindele, Hideo
Kigoshi (University of Tsukuba)
Modified DNA Aptamers that Bind to Vascular Endothelial Growth Factors
Naoto Honda, Kenta Hagiwara, Yuuya Kasahara, Masayasu Kuwahara
(Gunma University)
Exploring the Interaction of Quantum Dots Hybrid with Cells
San Kyeong, Cheolhwan Jeong, Bong-Hyun Jun, Yoon-Sik Lee
(Seoul National University)
Fluorescence Probe Development for Lung Cancer Stem Cells
Yong-An Lee, Srikanta Sahu, Animesh Samanta, Hwa-Young Kwon, Nam-Young
Kang, Young-Tae Chang (Singapore Bioimaging Consortium)
APEX-Generated ‘Barcode’ Tells Spatial Information of Target Protein in Living
Mammalian Cells
Song-Yi Lee, Hyun Woo Rhee (Ulsan National Institute of Science and Technology)
P-33
Proteome reactivity profiling for the discrimination of pathogenic bacteria
Jun-Seok Lee (Korea Institute of Science and Technology)
P-34
A Facile Method for Sequence Determination of Cyclic Peptides/Peptoids via CNBrMediated One-Pot Ring-Opening/Cleavage Reaction
Kang Ju Lee, Hyun-Suk Lim (Pohang University of Science and Technology)
P-35
Layer-by-Layer-Based Silica Encapsulation of Individual Yeast
Hojae Lee, Daewha Hong, Taegyun Park, Hee Chul Moon, Insung S. Choi
(Korea Advanced Institute of Science and Technology)
P-36
Dual-labeled multivalent glycoconjugates for detection of cell-surface lectins
Hui Li, Xizhe Tian, Kyung-Hwa Baek, Injae Shin (Yonsei University)
P-37
Structural Basis Underlying the Binding Specificity of Human Galectins-1, -3 and -7
for Galβ1-3/4GlcNAc
Tung-Ju Hsieh, Hsien-Ya Lin, Chuen-Jiuan Huang, Chun-Hung Lin (Academia Sinica)
P-38
DNA structure and stability under molecular crowding conditions
with cationic polymers
Daisuke Miyoshi, Yu-mi Ueda, Naohiko Shimada, Shu-ichi Nakano, Naoki Sugimoto,
Atushi Maruyama (Konan University)
P-39
Design, Synthesis, and Evaluation of Phenyl-Piperazine-Triazine-Based α-Helix
Mimetics Targeting Protein-Protein Interactions
Heejo Moon, Woo-Sirl Lee, Misook Oh, Hyun-Suk Lim
(Pohang University of Science and Technology)
- 13 -
P-40
P-41
P-42
P-43
P-44
P-45
P-46
P-47
P-48
P-49
P-50
P-51
Curvature Sensitive Membrane Perturbation by Adenovirus-derived Amphypathic
Peptides
Tomo Murayama, Sílvia Pujals, Shiroh Futaki (Kyoto University)
Development of the Strategy for the Selective Chemical Modification in an Abasic
Site of Duplex DNA
Norihiro Sato, Gen-ichiro Tsuji, Kazumitsu Onizuka, Fumi Nagatsugi
(Tohoku University)
Solution structure of P450 from Bacillus megaterium by small-angle X-ray
scattering
Ji-Hye Oh, Donghyuk Shin, Seungsu Han, Tae Yong Jee, Sei Young Lee, Yang-Gyun
Kim, Sangho Lee (Sungkyunkwan University)
Assembled, Mid-sized Agents that Modulate Protein-protein Interactions
Mai Tsubamoto, Prakash Parvatkar, Louvy Punzalan, Jiashi Sun, Nobuo Kato,
Motonari Uesugi, Junko Ohkanda (Kyoto University)
Artificial Glutamate Analogs as a Ligand for Neuronal Receptors
Masato Oikawa, Manami Chiba (Yokohama City University)
Design of Highly Reactive Peptide Tag for Protein Labeling and Protein Functional
Analysis
Shigekazu Tabata, Hirokazu Fuchida, Munetsugu Kido, Itaru Hamachi, Ryuichi
Shigemoto, Akio Ojida (Kyushu University)
Peptide microarrays for rapid analysis of peptide-RNA interactions
Sookil Park, Jaeyoung Pai, Injae Shin (Yonsei University)
Ppa-iRGD-Q as a Facile Depot Forming Peptide for Effective Photodynamic Therapy
Sung Jun Park, Hong-Jun Cho, Sehoon Kim, Yoon-Sik Lee (Seoul National University)
Design and Synthesis of Molecules Targeting the Non-enzymatic Tau protein in
cells
Tingting Chu, Tian Qiu, Yanmei Li (Tsinghua University)
Identification of Sub-Mitochondrion Proteome in Living Mammalian Cells By Using
Engineered Ascorbate Peroxidase (APEX)
Hyun Woo Rhee (Ulsan National Institute of Science and Technology)
Covalent labeling and real-time imaging of cytokine receptors on cell membrane
using DMAP-tethered cytokines
Shohei Uchinomiya, Yosuke Takaoka, Takahiro Hayashi, Yoshiaki Fukuyama, Itaru
Hamachi (National University of Singapore)
In vitro selection of macrocyclic peptides that trap the open state structure of
channelrhodopsin
Xiao Song, Hideaki E. Kato, Andrés D. Maturana, Takayuki Katoh, Yuki Goto,
Osamu Nureki, Hiroaki Suga (The University of Tokyo)
P-52
A Chemical Fluorescent Probe for the Detection of Aβ Oligomers.
Chai Lean Teoh, Dongdong Su, Srikanta Sahu, Seong-Wook Yun, Young-Tae Chang
(Singapore Bioimaging Consortium)
P-53
Crystallographic studies of adenylate kinase from Streptococcus pneumoniae D39
in new conformations
Trung Thanh Thach, Sangho Lee (Sungkyunkwan University)
- 14 -
P-54
Fabrication of Self-Assembling Peptide Hydrogels from Short Designed Peptides for
Cell Culture
Hiroshi Tsutsumi, Hisakazu Mihara (Tokyo Institute of Technology)
P-55
Luminescent Graphene Oxide/Peptide-Quencher Hybrids for Optical Detection of
Cell-Secreted Proteases by Turn-on Response
Jin-Kyoung Yang, Seon-Yeong Kwak, Su-Ji Jeon, Hye-In Kim, Joonhyuk Yim, Homan
Kang, San Kyeong, Jong-Ho Kim, Yoon-Sik Lee (Seoul National University)
P-56
Study on the Novel Chemical Probes to Analyze Protein-ligand Interactions
Kozo Yoneda, Yaping Hu, Masaki Kita, Hideo Kigoshi (University of Tsukuba)
P-57
Development of Highly Potent Macrocyclic Peptide Inhibitors Targeting Cofactorindependent Phosphoglycerate Mutase
Hao Yu, Patricia Dranchak, James Inglese, Hiroaki Suga (the University of Tokyo)
P-58
Mitochondria and Lysosome targetable Two-Photon Fluorescent Probes for
Hypochlorous Acid Detection and Imaging in Live Cells and Tissues
Lin Yuan, Lu Wang, Sung-Jin Park, Keshari Bikram, Juanjuan Peng,
Balasubramaniam Sivaraman, Young-Tae Chang (National University of Singapore)
- 15 -
Invited Speaker Abstracts
- 16 -
Deprotection chemistry-mediated protein activation in living cells
Peng Chen1,2
Beijing National Laboratory for Molecular Sciences & Molecular Engineering
Synthetic and Functional Biomolecules Center,
Peking University, Beijing 100871, China. 1
Peking-Tsinghua Center for Life Sciences, Beijing, China.2
pengchen@pku.edu.cn
Employing small molecules or chemical reagents to modulate the function of an intracellular
protein of interest, particularly in a gain-of-function fashion, remains a challenge. In this talk,
I will briefly introduce a “chemical uncaging” strategy that relies on a palladium-mediated
deprotection reaction to control protein activation in living cells. We identified biocompatible
and efficient palladium catalysts for propargylcarbamate cleavage, which allowed the
liberation of a free lysine from its propargyloxycarbonyl-protected lysine analogue that was
genetically and site-specifically incorporated into proteins. This “bioorthogonal protection
group-catalyst pair” was utilized for caging and subsequent releasing of a crucial lysine
residue from a bacterial TypeIII effector protein that modulates MAPK signaling pathway
within host cells. Our strategy extended the rapidly emerging palladium-mediated intracellular
chemistry from small molecules to proteins, which may be generally applicable for chemically
rescuing an essential lysine residue from a given protein, thus manipulating its activity within
a native cellular context.
Reference
Li J, et al. Nat. Chem. 2014, 6, 352-61.
Yang M, et al. Chem. Soc. Rev. 2014, DOI:10.1039/C4CS00117F
Peng Chen obtained his BS degree in Chemistry from Peking University
in 2002 and Ph.D with Prof. Chuan He in Chemistry at The University of
Chicago in 2007. After a postdoctoral training with Prof. Peter Schultz at
The Scripps Research Institute between 2007 and 2009, he started his
independent career as an Investigator in Chemical Biology at Peking
University in July 2009. His Lab is interested in developing and applying
novel chemistry tools to investigate protein-based interactions and
activities in living cells. He received NSFC Distinguished Young Scholar
Award in 2012, China Young Scientists Award in 2013 and Chemical
Society Review Emerging Investigator lectureship in 2014.
- 17 -
Turn-ON fluorescent affinity labeling using a small
bifunctional O-nitrobenzoxadiazole unit
Mikiko Sodeoka1,2
1
Synthetic Organic Chemistry Laboratory and CSRS, RIKEN, Japan
2
ERATO and CREST, JST, Japan
sodeoka@riken.jp
Affinity labeling has become a powerful tool to identify target proteins, as well as to
visualize/characterize target functions in living cells. However, although various functional
groups have been utilized for affinity labeling, new functional groups having better profile are
still demanded. To address this issue, we designed a simple chemical probe bearing a small
bifunctional O-NBD unit (NBD: nitrobenzoxadiazole). O-NBD unit selectively reacted with
amino group of the lysine residue to give the fluorescent N-NBD unit. Model ligand-protein
experiments showed that the O-NBD unit has excellent characteristics for target-specific
labeling even in the presence of a large excess of non-target proteins. Moreover, attachment
of the O-NBD unit to N,N-dialkyl-2-phenylindol-3-ylglyoxylamides (PIGAs), which are recently
developed translocator protein (TSPO) ligands, enabled us to visualize mitochondria
expressing TSPO in living cells by means of “turn-ON” fluorescence.
Reference
Takao Yamaguchi, Miwako Asanuma, Shuichi Nakanishi, Yohei Saito, Masateru Okazaki,
Kosuke Dodo and Mikiko Sodeoka, Chem. Sci. 5, 1021-1029 (2014).
Mikiko Sodeoka received her B.S. (1981), M.S. (1983) and Ph.D. degree
(1989) from Chiba University. After working at Sagami Chemical
Research Center (1983-1986), she joined the Faculty of Pharmaceutical
Sciences, Hokkaido University as a research associate. After working as
a post-doctoral fellow at Harvard University, she moved to the University
of Tokyo (1992). She became a group leader at Sagami Chemical
Research Center in 1996 and an associate professor of the University of
Tokyo in 1999. In 2000, she moved to Tohoku University as a full
professor. Since 2006 she has been the chief scientist of the Synthetic
Organic Chemistry Laboratory at RIKEN.
- 18 -
Detection of native proteins in live cells by iFIT (intrinsic Förster resonance
energy transfer Imaging Technology)
Sang J. Chung
Department of Chemistry Dongguk University, Seoul, Korea
sjchung@dongguk.edu
Using tryptophan fluorescence as a FRET donor, we have developed a label-free protein
imaging system, so called intrinsic fluorescence resonance energy transfer (iFRET) imaging
technology (iFIT), to enable a specific detection of target proteins in live cells. In iFRET, the
probes bind the target proteins in a specific manner to accept Förster resonance energy from
the tryptophan residues near the binding site, resulting in a strong FRET. By developing
good iFRET acceptor fluorescence moieties and a deep UV microscope, we have succeeded
to detect target proteins in live cells without any modification thereof. This label-free method
can be applied to detect specific proteins or screen drugs in live cells without any
modification on the target proteins. In this conference, the principle and the live cell imaging
results using iFIT will be presented.
References
Kim JH, et al. Mol. Biosystems 2014,10: 30-33.
Kang HJ, et al. submitted.
Sang Jeon Chung received his Ph.D. in Chemistry from POSTECH,
Korea, in 1996. He did his postdoctoral work with Prof. Chi-Huey Wong
at The Scripps Research Institute and Prof. Gregory Verdine at Harvard
University. In 2003, he was appointed a Senior Scientist at KRIBB and
promoted to Principal Scientist in 2009. In March 2013, he moved to
Dongguk University, Seoul, Korea. Currently he is an Associate
Professor of Chemistry Department and director of Molecular Targeting
Research Center of Dongguk University and also running Lab of
Chemical Biology at Dongguk University.
- 19 -
Nano- and Quantum-biodevices for Biomedical Applications
Yoshinobu Baba
Department of Applied Chemistry, School of Engineering, FIRST Research Center for Innovative
Nanobiodevices, Nagoya University
National Institute of Advanced Industrial Science and Technology (AIST)
babaymtt@apchem.nagoya-u.ac.jp
Nano-/quantum-biodevice is a piece of contrivance, equipment, machine, or component,
which is created by the overlapping multidisciplinary activities associated with nano/quantum-technology and biotechnology, intended for biological, medical, and clinical
purposes. In this lecture, I will describe the development of nano-/quantum-biodevices for
biomedical applications, including single cancer cell diagnosis for cancer metastasis,
circulating tumor cell (CTC) detection by microfluidic devices, nanopillar devices for ultrafast
analysis of genomic DNA and microRNA, nanopore devices for single DNA and microRNA
sequencing, nanowire devices for exosome analysis, single-molecular epigenetic analysis,
quantum switching in vivo imaging of iPS cells and stem cells, and quantum technologybased cancer theranostics [1-10]. Euglena-based “biomimetic mechanical system” enables
us to develop reliable circulating tumor cell (CTC) separation and detection technique for
cancer metastasis diagnosis. Immunopillar devices realized the fast and low invasive “from
blood to analysis” type biomarker detection of cancer with fM detection sensitivity within 2
min. Additionally, nanopillar devices give us ultrafast separation of DNA and microRNA
within 60 µs and nanopillar-nanopore integrated nanobiodevice enables us ultarafast single
molecular DNA sequencing. Nanowire devices coupled with super-resolution optical
microscopy are extremely useful to analyze exosomes from cancer cells and exosomal
microRNA analysis. Quantum dots are applied to develop nanobiodevice for single cancer
cell diagnosis, single molecular epigenetic analysis, quantum switching in vivo imaging for
iPS cell and stem cell therapy and theranostic device for cancer diagnosis/therapy.
References
1. N. Kaji, Y. Baba, et al., Chem. Soc. Rev., 39, 948 (2010).
2. M. Tabuchi, Y. Baba, et al., Nature Biotech., 22, 337 (2004).
3. R. Bakalova, Y. Baba, et al., Nature Biotech., 22, 1360 (2004).
4. Y.S. Park, Y. Baba, et al., ACS Nano., 4, 121 (2010).
5. M. F. Serag, Y. Baba, et al., ACS Nano., 5, 493, (2011).
6. T. Yasui, Y. Baba, et al., ACS Nano, 5, 7775 (2011).
7. M.F. Serag, Y. Baba, et al., Nano Lett., 12, 6145 (2012).
8. T. Yasui, Y. Baba, et al., ACS Nano, 7, 3029 (2013).
9. K. Hirano, Y. Baba, et al., Nano Lett., 13, 1877 (2013).
10. J. Wang, Y. Baba, et al., Biosens. Bioelec., in press (2014).
Yoshinobu Baba, who received his Ph.D. in chemistry from Kyushu
University in 1986, is a professor at Department of Applied Chemistry,
Graduate School of Engineering, Nagoya University. He is also a
Director of FIRST Research Center for Innovative Nanobiodevices,
Nagoya University. He is an Associate Editor of Anal. Chem. and
serving to over 20 scientific journals as an editorial board member. He
received over 100 awards for his contributions in nanobiotechnology.
His major area of interest is nanobiosicence and nanobiotechnology for
medical diagnosis, tissue engineering, and molecular imaging. He is the
author or co-author of 798 publications, including research papers,
proceedings, reviews, and books. He has delivered more than 780 plenary and invited
lectures at conferences. His work has been cited on 309 occasions by newspapers and
televisions.
- 20 -
Cellular and in vivo Imaging Probes with Tunable Chemical Switches
Kazuya Kikuchi1,2
1
2
Graduate School of Engineering, Osaka University, Japan
WPI-Immunology Frontier Research Center, Osaka University, Japan
kkikuchi@mls.eng.osaka-u.ac.jp
One of the great challenges in the post-genome era is to clarify the biological significance of
intracellular molecules directly in living cells. If we can visualize a molecule in action, it is
possible to acquire biological information, which is unavailable if we deal with cell
homogenates. One possible approach is to design and synthesize chemical probes that can
convert biological information to chemical output.
Fluorescence protein labeling by synthetic probes is a powerful approach to investigate
protein function and localization inside living cells. This chemistry-based technique utilizes a
pair of a protein tag and its specific ligands connected to fluorophores. Its potential
advantage is that various fluorescent molecules are available as labeling reagents, and the
timing of protein labeling is easily controlled. Because of these characteristics, this method is
attracting attention as an alternative of fluorescent proteins. We developed two fluorogenic
labeling systems, photoactive yellow protein (PYP) tag and mutant beta-lactamase (BL) tag.
Reference
Mizukami S, et al. Acc Chem Res. 2013,47:247-256.
Kazuya Kikuchi received his Ph.D. in Pharmaceutical Sciences from The
University of Tokyo, Japan, in 1994. He did his postdoctoral work with
Prof. Roger Y Tsien at UC Berkeley and with Prof. Donald Hilvert at The
Scripps Research Institute. In 1997, he was appointed Assistant
Professor at The University of Tokyo and promoted to Associated
Professor in 2000. In 2005, he moved to Osaka University as a full
professor.
- 21 -
Integrative chemical biology approaches to examine histone posttranslational
modifications
Xiang David Li
Department of Chemistry, The University of Hong Kong, Hong Kong
xiangli@hku.hk
Histone posttranslational modifications (PTMs), such as phosphorylation, methylation and
acetylation, play crucial roles in regulating many fundamental cellular processes, such as
gene transcription, DNA replication, DNA damage repair, chromosome segregation and cell
differentiation. Increasing evidences have indicated that PTMs of histones can serve as a
heritable „code‟ (so-called „histone code‟), which provides epigenetic information that a
mother cell can pass to its daughters. Histone code is „written‟ or „erased‟ by enzymes that
generate or remove the modifications of histones. Meanwhile, „readers‟ of histone code
recognize specific histone modifications and „translate‟ the code by executing distinct cellular
programs necessary to establish the diverse cell phenotypes, while the genetic code (DNA)
is unaltered.
While a large number of PTMs have been identified on various sites of histones, so far only a
handful of them have been extensively studied. The cellular mechanisms and functions of
many other PTMs, particularly those newly identified ones, remain essentially obscure. In this
talk, I will present our chemical biology approaches to unravel biological significance of
histone PTMs by identifying their substrates, „writers‟, „erasers‟ and „readers‟.
Xiang David Li, born 1981 in Kunming, China, received his B.Sc. in
Chemistry at Fudan University in 2003, and Ph.D. in 2008 from The
University of Hong Kong under the guidance of Professor Dan Yang. He
has then spent three years as a postdoctoral fellow with Professor Tarun
Kapoor at Rockefeller University. In 2011, he joined the University of
Hong Kong as an Assistant Professor of Chemistry. His research
interest is to develop chemical approaches to study protein
posttranslational modifications.
- 22 -
Design and Construction of Fluorescent Nucleic Acid Systems
Byeang Hyean Kim
Department of Chemistry, Pohang University of Science and Technology (POSTECH),
790-784, Pohang, Korea
bhkim@postech.ac.kr
Sequence specific hybridization probes composed of fluorescent synthetic oligonucleotides
have been developed for genetic analysis in the post-genomic era. Currently such
fluorescent oligonucleotides play a decisive role in analysis of the genetic information and
DNA sensing such as SNP typing. 1
Fluorescent nucleic acid systems are widely applied in various fields, from fundamental
biological probes to nano-construction, 2 Nucleic acids are used as a scaffold for arranging
aromatic fluorophore assemblies, either by insertion into the DNA base pairs or by stacking
via the duplex. Moreover, chemical modifications of nucleic acids are accessible by the
modified DNA phosphoramidites or postsynthetic approach, and provide with new and
interesting fluorescent nucleic acids systems. Fluorescent nucleic acid systems represent an
extensive and exciting research area in chemistry as well as in biotechnology and
photophysics. We have synthesized and investigated new fluorescent nucleic acid systems
for probing single nucleotide polymorphisms (SNPs),3 structural changes of DNA4 and ligand
interaction with RNA bulge.
We have developed the new type of molecular beacon, quencher-free molecular beacon
(QF-MB), that exhibits several advantageous features, including a high level discrimination
between the target and its single-mismatched congeners and an economical device set-up
due to the absence of the quencher. We have also designed and synthesized the probing
system for quadruplex structures of DNA (G-quadruplex and i-motif) and B-Z transition.
Strong π–π stacking interactions in nucleic acids can be used to generate novel secondary
structures. We have investigated the fluorescent phenomena and structures of pyrene
modified oligodeoxyadenylate and oligodeoxyguanylate. The covalently linked pyrenes
induced the formation of a self-assembled olgiodeoxyadenylate duplex5 and various
secondary structures with interesting fluorescence phenomena.6
References
1. Lee, I. J., Kim, B. H., Labeling oligonucleotides toward the biomedical probe, in Medicinal
Chemistry of Nucleic Acids, Zhang, L-H., Xi, Z., J. Chattopadhyaya Eds., Hoboken, NJ,
Wiley, 2011, 292-334.
2. Venkatesan, N., Seo, Y. J., Kim, B. H., Chem. Soc. Rev., 2008, 37, 648-663.
3. Hwang, G. T., Seo, Y. J., Kim, B. H., J. Am. Chem. Soc., 2004, 126, 6528-6529.
4. Lee, I. J., Yi, J. W., Kim, B. H., Chem. Commun., 2009, 36, 5383-5385.
5. Seo, Y. J., Rhee, Y. M., Joo, T., Kim, B. H., J. Am. Chem. Soc., 2007, 129, 5244-5247.
6. (a) Park, J. W., Seo, Y. J., Kim, B. H., Chem. Commun., 2014, 50, 52-54. (b) Kim, K. T.,
Veedu, R. N., Seo, Y. J., Kim, B. H., Chem. Commun., 2014, 50, 1561-1563
B. H. Kim got his undergraduate education at Seoul National University
and received his Ph.D. from University of Pittsburgh in 1987. He worked
as a postdoctoral associate with Professor K. C. Nicolaou at the
University of Pennsylvania from 1987 to 1988. Then he returned to his
home country to become an Assistant Professor of Pohang University of
Science and Technology (POSTECH). He has served as Head of the
Department of Chemistry and is now Professor of Chemistry at
POSTECH.
- 23 -
Structures, Interactions, and Functions of Nucleic Acids
under Molecular Crowding Conditions
Naoki Sugimoto1,2*
Frontier Institute for Biomolecular Engineering Research (FIBER), and 2Graduate School of Frontiers
of Innovative Research in Science and Technology (FIRST), Konan University, Kobe, Japan
sugimoto@konan-u.ac.jp
How does molecular crowding affect the stability of nucleic acid structures inside cells?
Water is the major solvent component in living cells, and the properties of water in the highly
crowded media inside cells differ from that in buffered solution. As it is difficult to measure
the thermodynamic behavior of nucleic acids in cells directly and quantitatively, we recently
developed a cell-mimicking system using cosolutes as crowding reagents. In this
presentation, I will show how the structures and thermodynamic properties of nucleic acids
differ under various conditions such as highly crowded environments and discuss the major
determinants of the crowding effects on nucleic acids. The effects of molecular crowding on
noncanonical structures of DNA and RNA quadruplexes that play important roles in
transcription and translation are also discussed.
Acknowledgements: The author is grateful to the colleagues named in the cited papers from my laboratory
and institute (FIBER), especially Drs Endo, Tateishi-Karimata, Takahashi, Nakano, Miyoshi, Nagatoishi,
Kuwahara, Pramanik, Rode, and my technical assistant Ms. Yamaguchi. This work was partly supported by the
Grants-in-Aid for Scientific Research, the Ministry of Education, Culture, Sports, Science and Technology
(MEXT), Japan and MEXT Program for the Strategic Research Foundation at Private Universities, the Nagase
Science and Technology Foundation, the Hirao Taro Foundation of the Konan University Association for
Academic Research, and Chubei Itoh Foundation.
References
Sugimoto and colleagues, J. Am. Chem. Soc., 2006, 128, 7957-7963; J. Am. Chem. Soc., 2006, 128,
15461-15468; J. Am. Chem. Soc., 2007, 129, 5919-5925; Angew. Chem. Int. Ed., 2008, 47, 9034-9038;
J. Am. Chem. Soc., 2009, 131, 3522-3531; J. Am. Chem. Soc., 2009, 131, 9268-9280l; J. Am. Chem.
Soc., 2009, 131, 16881-16888; Biochemisry, 2010, 49, 4554-4563; Biochemistry, 2011, 50, 7414–
7425; Angew. Chem. Int. Ed., 2012, 51, 1416-1419; J. Am. Chem. Soc., 2013, 135, 9412-9419; Angew.
Chem. Int. Ed., 2013, 52, 5522-5526; Nucleic Acids Res., 2013, 41, 6222-6231; Angew. Chem. Int. Ed,,
2013, 52, 13774-13778; International Review of Cell and Molecular Biology (Elsevier), 2014, 307,
Chap. 8, 205-273l; Chem. Rev., 2014, 114, 2733-2758; Sci. Rep., 2014, 4, 3593; PLoS ONE, 2014,
9(3): e90580; ); J. Phys. Chem. B., 2014, 118, 379-389; Nucleic Acids Res., 2014, in press and so on.
Naoki Sugimoto received Ph.D. in 1985 from Kyoto University, Japan. After
postdoctoral work at University of Rochester in USA, he joined Konan
University, Kobe, Japan in 1988 and is a full professor since 1994. From 2003,
he holds a director of Frontier Institute for Biomolecular Engineering Research
(FIBER) at Konan University. He is a member of the Editorial Board of Nucleic
Acids Research from 2007 to the present, a first chairman of Forum on
Biomolecular Chemistry (FBC) from 1998 to 2001, and a chairman of Division of
Biofunctional Chemistry of the Chemical Society of Japan (CSJ) from 2011 to
2013. He received the Dr. Masao Horiba‟s Award in 2004, Distinguised Scientist
Award from ICA (International Copper Association), New York, USA in 2005,
Hyogo Science Award from Hyogo Prefecture, Japan in 2006, the CSJ Award for Creative Work in
2007, and so on. His research interests focus on Biofunctional Chemistry, Biomaterials, Bio-Nano
Engineering, Molecular Design, Biofunctional Chemisty, and Biotechnology. He has published about
500 scientific papers, reviews, and books.
- 24 -
Distinct role of water in protein-protein interactions
Sihyun Ham
Department of Chemistry, Sookmyung Women’s University
Hyochangwon-gil 52, Yongsan-gu, Seoul 140-742, Korea
sihyun@sm.ac.kr
Understanding the molecular determinants of the relative propensity for proteins to aggregate
in a cellular environment has been a central issue in attacking protein-aggregation diseases
and in the development of human therapeutics. Despite the expectation that the protein
aggregation can largely be attributed to the direct protein-protein interactions within an
aggregate or in solution, we here unveil a crucial role of hydration water in ruling the
aggregation propensity of proteins both in vitro and in vivo. The protein overall hydrophobicity,
defined solely by the hydration free energy of a protein in its monomeric state sampling its
equilibrium structures, was shown to predominantly dictate the protein aggregation
propensity in aqueous solutions. We also find striking discrimination by the hydration water of
positively and negatively charged residues depending on the protein net charge in regulating
the solubility of a protein, which establishes novel design strategies for the biotechnological
generation of aggregation-resistant proteins as biotherapeutics.
References
1. Chong, Ham, Angew. Chem. Int. Ed. Engl., 53, 3961, (2014).
2. Chong, Ham, Proc. Natl. Acad. Sci. USA, 109, 7636, (2012).
3. Chong, Lee, Kang, Park, Ham, J. Am. Chem. Soc. 133, 7075, (2011).
Sihyun Ham received her Ph.D. in Chemistry from Texas Tech University,
USA, in 1998. She did her postdoctoral work with Prof. Valerie Daggett
at Univ. of Washington, Seattle from 1998-2000. She was appointed as a
research professor at Korea University from 2001 to 2003. In 2003 fall,
she joined as a faculty of Chemistry department, Sookmyung Women‟s
Univ in Seoul, Korea. Currently she is a full professor of Chemistry and
running Nano/Bio computational Lab. in Sookmyung.
- 25 -
Metalloprotein design using genetic code expansion
Jiangyun Wang
Institute of Biophysics, Chinese Academy of Sciences
15 Datun Road, Chaoyang District, 100101
jwang@ibp.ac.cn
One aim in our laboratory is to use small, soluble protein scaffold, and the genetic
incorporation of unnatural amino acid to design easy-to-characterize, easy-to-produce, and
easy-to-optimize metalloenzymes which catalyze these important reactions with equal or
greater efficiency/selectivity than that of the natural systems. Through the genetic
incorporation of the Tyr-His ligand and CuB site into myoglobin, we recapitulated important
features of HCO into this small soluble protein, which exhibits selective O2 reduction activity
while generating less than 6% ROS, at more than 1000 turnovers. These results support that
Tyr-His crosslink is indeed important for HCO function, and creates the exciting opportunity
to rapidly evolve better HCO model proteins to achieve higher activity and selectivity, which
may be suitable as alternatives to precious metal catalyst in fuel cells.
Another aspect of our ongoing research is the development of new methods for precise
attachment of functional metal complexes on biomolecules, which is an important strategy for
metalloprotein design. Bioorthogonal chemical reactions together with genetic code
expansion technique have provided exciting new means for protein labeling in living cells.
The main advantages of photoclick reaction are its fast rate (up to 50 M-1S-1), and that it has
no need for toxic copper catalyst.
References
1.
2.
3.
4.
5.
6.
Li, F. H.; Shi P.; Li J. S.;Yang F.; Wang T. Y.; Zhang W.; Gao F.; Ding W.; Li D.; Li J.; Xiong Y.; Sun J. P.;
Gong W. M.; Tian C. L.; Wang J. Y., “A Genetically Encoded 19F NMR Sensor for Tyrosine Phosphorylation
” Angew. Chem. Intl. Ed. 2013, 52, 3958-62.
Zhou Q.; Hu M. R.; Zhang W.; Jiang, L.; Perrett S.; Zhou J.; Wang J. Y., “Probing the Function of the Tyr-Cys
Crosslink in Metalloenzymes through the Genetic Incorporation of 3-Methylthiotyrosine” Angew. Chem. Intl.
Ed. 2013, 52, 1203-7.
Liu, X. H.; Yu, Y.; Hu, C.; Zhang, W.; Lu, Y.; Wang, J. Y, “Significant Increase of Oxidase Activity through the
Genetic Incorporation of a Tyrosine-Histidine Cross-Link in a Myoglobin Model of Heme-Copper Oxidase”
Angew. Chem. Intl. Ed. 2012, 51 , 4312-6. (News of the Week, Chemical and Engineering News)
Liu, X. H.; Li, J. S.; Dong, J. S.; Hu , C.; Gong, W. M.; Wang, J. Y., “Genetic Incorporation of a Metal
Chelating Amino Acid as a Probe for Protein Electron Transfer” Angew. Chem. Intl. Ed. 2012, 51, 10261-5.
(Very Important Paper and Cover Article)
Yu, Z.; Pan, Y. C.; Wang. Z. Y.; Wang, J. Y.; Lin. Q., “Genetically Encoded Cyclopropene Directs Rapid,
Photoclick Chemistry- Mediated Protein Labeling in Mammalian Cells” Angew. Chem. Intl. Ed. 2012, 51，
10600-4.
Wang, J. Y.; Zhang, W.; Song, W. J.; Wang, Y. Z.; Yu, Z. P.; Li, J. S.; Wu, M. H.; Wang, L.; Zang, J. Y.; Lin,
Q., “A Biosynthetic Route to Photoclick Chemistry on Proteins” J. Am. Chem. Soc. 2010, 132, 14812-8.
Jiangyun Wang holds a diploma degree in Physical Chemistry from the
University of Science and Technology of China. He finished his PhD under
the supervision of Kenneth Suslick at the University of Illinois at UrbanaChampaign in 2003. Thereafter, he worked as a Research Fellow in the
Peter Schultz Group of the Scripps Research Institute in San Diego,
California. Since January 2008 he is head of the Institute of Biophysics
Group for Chemical Biology. His present research interests includes:
1.
Genetic code expansion to facilitate protein labeling and
engineering in various host organisms, including bacteria cyanobacteria,
mammals and plants.
2.
Development of new bio-orthogonal click chemical reactions to
facilitate protein and RNA labeling, as wells as super-resolution imaging in
living cells.
3. Design of metalloproteins, which includes cytochrome c oxidase, nitrogenase, photosystem II.
- 26 -
Towards Pharmacological Modulation of Protein-Protein Interactions
Hyun-Suk Lim1,2
1
Department of Chemistry, Pohang University of Science & Technology, South Korea
Department of Biochemistry & Molecular Biology, Indiana University School of Medicine,
Indianapolis, Indiana, USA
hslim@postech.ac.kr
2
The majority of drugs on the market today target proteins with defined small-molecule
binding sites, including enzymes and receptors. However, some of the most pressing and
devastating diseases involve proteins that do not possess these natural binding sites, such
as those involved in protein-protein interactions (PPIs) associated with many cancers, and
protein-misfolding, associated with most neurodegenerative diseases. Thus, molecules that
can modulate PPIs or protein stability would be valuable research tools to uncover molecular
functions of target proteins and could be potential therapeutic candidates. However,
designing or discovering such molecules is challenging because of the relatively featureless
protein surfaces, the lack of suitable chemical libraries, and the shortage of efficient highthroughput screening methods. In this presentation, I will present our attempts to address
these challenges by developing novel types of synthetic molecules that can mimic protein
surface structures, and establishing a facile high-throughput screening method that allows for
rapid and efficient discovery of potential pharmacological modulators of PPIs.
Hyun-Suk Lim received his Ph.D. in Chemistry from POSTECH, Korea
in 2004. He did his postdoctoral work with Prof. Thomas Kodadek at UTSouthwestern Medical Center at Dallas. In 2008, he was appointed
Assistant Professor at Indiana University School of Medicine. In 2012,
he moved to POSTECH. Currently he is an Associate Professor of
Chemistry Department of POSTECH.
- 27 -
Peptide-based approaches for delivering exogenous molecules into cells
Shiroh Futaki
Institute for Chemical Research, Kyoto Univeristy, Uji, Kyoto 611-0011, Japan
futaki@scl.kyoto-u.ac.jp
Arginine-rich cell-penetrating peptides, including octaarginine (R8) and HIV-1 TAT peptides,
have the ability to transport exogenous bioactive molecules into cells [1]. Endocytosis, a
physiological engulfing system of extracellular molecules/liquids into cells plays a significant
role in this transport [2]. However, a dramatic change in the methods of internalization for
these peptides is brought about by the presence of pyrenebutyrate, a counteranion bearing
an aromatic hydrophobic moiety [3,4]. The addition of pyrenebutyrate results in direct
membrane translocation of the peptides yielding diffuse cytosolic peptide distribution within a
few minutes. Using this method, rapid and efficient cytosolic delivery of the enhanced green
fluorescent protein (EGFP) was achieved in cells including rat hippocampal primary cultured
neurons. We next explored the effects of PyB on the promotion of R8 translocation using
model membranes having liquid-ordered (Lo) and liquid-disordered (Ld) phases [5]. Confocal
microscopic observations of giant unilamellar vesicles (GUVs) showed that PyB significantly
accelerated the accumulation of R8 on membranes containing negatively charged lipids,
leading to the internalization of R8 without collapse of the GUV structures. PyB displayed an
alternative activity, increasing the fluidity of the negatively charged membranes, which
diminished the distinct Lo/Ld phase separation on GUVs. This was supported by the
decrease in fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene (DPH). Additionally,
PyB induced membrane curvature, which has been suggested as a possible mechanism of
membrane translocation for R8. Taken together, our results indicate that PyB may have
multiple effects that promote R8 translocation through cell membranes.
References
1. I. Nakase et al. Acc. Chem. Res. 45, 1132 (2012)
2. S. Futaki et al., Curr. Pharm. Des. 19, 2863 (2013)
3. F. Perret et al, J. Am. Chem. Soc. 127, 1114 (2005)
4. T. Takeuchi et al., ACS Chem. Biol. 1, 299 (2006)
5. S. Katayama et al., Biochim. Biophys. Acta. 1828, 2134 (2013)
Shiroh Futaki obtained his Ph.D. in 1989 from Kyoto University, Japan.
Following his appointment as a Research Associate and an Associate
Professor at the University of Tokushima, he moved to Kyoto University
in 1997. Meanwhile, he spent 16 months (1989-1991) in the US as a
Postdoctoral Associate at Rockefeller University (James Manning
laboratory). He has been a Professor of Biochemistry at the Institute of
Chemical Research, Kyoto University, since 2005. His research interests
include design of bioactive peptides having unique functions (ionchannel formation, cell penetration, DNA-binding, and so on).
- 28 -
Biomedical Applications of a Novel Class of High-affinity Peptides (Aptides)
Sangyong Jon
KAIST Institute for the BioCentury, Department of Biological Sciences, Korea Advanced Institute of
Science and Technology (KAIST), 291 Daehak-ro, Daejeon 305-701, Korea
syjon@kaist.ac.kr
Aptides, a novel class of high-affinity peptides, are designed to possess a “tweezers-like”
structure comprising a unique structure-stabilizing scaffolding region and two high-affinity
target-binding components. We have produced aptides with nanomolar-range binding
affinities for several targets using phage display selection, demonstrating the utility of this
technology. Indeed, peptides with high target affinity isolated from aptide libraries have
offered the potential for use as diagnostic or drug candidates. An aptide targeting the tumor
biomarker fibronectin extradomain B (EDB) showed remarkable in vivo accumulation at the
tumor site. Thus we have utilized the anti-EDB aptide for various biomedical applications,
including targeted drug delivery, cancer imaging by MRI, and targeted protein therapeutics.
In this lecture, bio-inspired design, characterization, and a couple of biomedical applications
of aptides will be presented.
References
1. Kim S, et al. Angew. Chem. Int. Ed. 2012, 51:1890-1894.
2. Saw PE, et al. J. Mater. Chem. B. 2013, 1:4723-4726.
Dr. Sangyong Jon received his Ph.D. in 1999 from the Department of
Chemistry at Korea Advanced Institute of Science and Technology
(KAIST). He had experienced his postdoc career in the Department of
Chemical Engineering at M.I.T under the supervision of Dr. Robert
Langer. In 2004, he joined GIST as an Assistant Professor of Life
Sciences and promoted to a Professor in 2010. He moved to KAIST in
2012 and is currently a Professor in the Department of Biological
Sciences at the institute.
- 29 -
Chemically synthetic glycopeptide vaccines
Yan-Mei Li1
1
Key Lab of Bioorganic Phosphorus Chemistry & Chemical Biology (Ministry of Education)
Department of Chemistry Tsinghua University Beijing 100084, P.R.China
liym@tsinghua.edu.cn
Carbohydrates and glycopeptides which are expressed on carcinoma cells surface can be
used as the targets for the immunotherapy. However, these targets always have weak
immunogenicity and cannot induce robust immune response. Many efforts have been taken
to enhance the immunogenicity. For example, chemically synthetic vaccines were conjugated
with the foreign carrier proteins and these vaccines could induce strong immune response.
Besides, multi-component vaccines and nanoparticle vaccines were also synthesized.
In our studies, MUC1 glycopeptide, which is over-expressed and aberrantly glycosylated in
tumor cells, was chosen as the target antigen. Firstly, MUC1 glycopeptide was conjugated
with Bovine Serum Albumin (BSA) to improve the immunogenicity [1]. However, BSA is highly
immunogenic and can suppress immune reaction, the improved vaccines containing T-helper
cell epitopes were developed [2]. To our knowledge, Toll-like receptor 2 ligand Pam3Cys is a
potent immune stimulant. Therefore we developed a three-component chemically synthetic
vaccine [3]. To further improve the efficiency of vaccines, cluster effect based on the dendritic
lysine was introduced in the vaccines [4]. Besides, to utilize the size effect of the vaccines and
avoid the side-effect of the extra adjuvant, a self-assembly adjuvant-free vaccine was
developed [5]. All of these vaccines can stimulate the immune system to elicit high level of
antibodies against MUC1 glycopeptides.
References
1. Cai, H.; Huang, Z. H.; Shi, L.; Sun, Z. Y.; Zhao, Y. F.; Kunz, H.; Li, Y. M. Angew.
Chem. Int. Ed. 2012, 51, 1719-1723.
2. Cai, H.; Chen, M. S.; Sun, Z. Y.; Zhao, Y. F.; Kunz, H.; Li, Y. M. Angew. Chem. Int.
Ed. 2013, 52, 6106-6110.
3. Cai, H.; Sun, Z. Y.; Huang, Z. H.; Shi, L.; Zhao, Y. F.; Kunz, H.; Li, Y. M. Chem.-Eur.
J. 2013, 19, 1962-1970.
4. Cai, H.; Sun, Z. Y.; Chen, M. S.; Zhao, Y. F.; Kunz, H.; Li, Y. M. Angew. Chem. Int.
Ed. 2014, 53, 1699-1703.
5. Huang, Z. H.; Shi, L.; Ma, J. W.; Sun, Z. Y.; Cai, H.; Chen, Y. X.; Zhao, Y. F.; Li, Y. M.
J. Am. Chem. Soc. 2012, 134, 8730-8733..
Yan-Mei Li received her Ph.D. in Chemistry from Tsinghua University,
P.R.China, in 1992. She did her postdoctoral work with Prof. Herbert
Waldmann at University of Karlsruhe, Germany. In 1993, she was
appointed Associate Professor at Tsinghua University and promoted to
Professor in 1998. She is the Director of Key Lab of Bioorganic
Phosphorus Chemistry & Chemical Biology, Ministry of Education from
2004. Currently she is the Vice Chair in Department of Chemistry,
Tsinghua University.
- 30 -
Robust and reliable high-throughput screening of bead-based peptide libraries
Su Seong Lee, Jaehong Lim, Joo-Eun Jee, Liqian Gao, Jessica Oon,
Yong Siang Ong
Institute of Bioengineering and Nanotechnology, 31 Biopolis Way, The Nanos, Singapore 138669
sslee@ibn.a-star.edu.sg
Combinatorial one-bead-one-compound (OBOC) peptide libraries have been used for affinity
screening. This approach appears attractive in terms of easy generation of sequence
diversity and peptide structure using chemistry platform. However, there are only a few
successful cases reported to date. It is not as popular as alternative methods such as phage
display and aptamers. Many efforts have been dedicated to identify the exact problems with
the bead-based screening. One possible drawback should be related to various nonspecific
interactions, inherited from this bead-based approach. Among the nonspecific/undesired
interactions, the types of fluorescence dyes for the labelling of target proteins significantly
affect screening resulting in significant interference of false positive beads. Use of
zwitterionic dyes could solve this big issue. Besides this, peptide loading density on beads
and other factors will be discussed in the presentation. Our current applications of the beadbased screening will also be introduced.
Peptide library – 1day
Assay & Automatic bead sorting – 1 day
5-mer (2 g)
mix
Dye-labeled protein
Split
Titan 357
Fully automatic synthesis
Split-and-mix method
CNBr cleavage
- 1 min
1 ~ 2 day
MALDI sampling
MALDI-TOF/TOF
Semi-automatic
peptide sequencing
MS & MS/MS data
50 peptides/hr
Dr. Lee received his MSc (1994) and PhD (1997) in inorganic chemistry
from Seoul National University in Korea. He worked at R&D center of
Saehan Industries Inc. as senior research scientist for three years.
Thereafter, he joined Prof. Hyeon‟s laboratory at Seoul National
University as a postdoctoral fellow in 2000, and then Prof. Ying‟s
laboratory at MIT as a postdoctoral fellow in 2001. He joined IBN in 2003
as a Senior Research Scientist. He was promoted to a Principle
Research Scientist in 2006. His research interests include
heterogeneous catalyst, synthesis of novel nanomaterials, glycomimetics
and development of high affinity protein capture agents
- 31 -
Protein phosphorylation of methanoarchaeon Methanohalophilus
portucalensis FDF1T in methanogenesis and osmoadaptation
Shih-Hsiung Wu1,2
1
Institute of Biological Chemistry, Academia Sinica, Taipei, TAIWAN
2
Institute of Biochemical Sciences and Department of Chemistry,
National Taiwan University, Taipei, TAIWAN
Shwu@gate.sinica.edu.tw
Halophilic methanoarchaeon Methanohalophilus portucalensis FDF1T was used as
an archaeal model for phosphoproteomic studies. Based on the data of
phosphoproteimic analysis, several important discoveries were found: (1) Unlike
previous studies on archaeal phopshoproteomics, pHis and pAsp were identified on a
small portion of phosphorylated peptides which revealed that two-component
systems exist in this methanogen. This result is consistent with genomic analysis
which showed 40 genes of two-component systems. (2) Almost all enzymes involved
in methanogenesis and the biosynthesis of betaine were phosphorylated. It meant
that single-carbon metabolism including methane generation and response to salt
stress was regulated by protein phosphorylation. In addition, some enzymes in
gluconeogenesis for biosynthesis of glucose from CO2 fixation are also
phosphorylated. (3) Many proteins participating in protein biosynthesis, folding and
degradation such as ribosome proteins, GroEL/GroES and proteaosome were
phosphorylated. It implied that protein metabolism was also significantly controlled by
protein phosphorylation. (4) Both of 21st and 22nd animo acids, selenocysteine and
pyrrolysine, were found in this methanogen and both selenocysteinyl and pyrrolysyl-tRNA synthases were phosphorylated. It might point out that the incorporation of both
uncommon amino acids was manipulated by protein phosphorylation. (5) Glycinesarcosine methyltransferase was phosphorylated in many sites and its activity was
affected by protein phosphorylation, particluarly in Tyr 169. Overall, in comparison
with eubacteria and other archaea, the protein phosphorylation in Methanohalophilus
portucalensis FDF1T was involved not only in carbohydrate and protein metabolism
but also in environmental adaptation such as osmotic stress and anaerobic survival.
Shih-Hsiung Wu is a Distinguished Research Fellow of the Institute of
Biological Chemistry, Academia Sinica, Taiwan. He received his Ph.D. in
1987 from University of Wisconsin, Madison (Prof. Charles J. Sih). He
became Associate Research Fellow (1988–1991) at Academia Sinica. In
1991, he was promoted to Research Fellow (1991-2008) and became a
Distinguished Research Fellow since 2008 His research interest focuses
on the structure-functional relationship of biomolecules including
polyketides, glycoconjugates, polysaccharides, peptides and proteins.
- 32 -
Development of programmable biological functionalities
Matthew W. Chang1,2
1
Department of Biochemistry and 2NUS Synthetic Biology Research Consortium,
National University of Singapore, Singapore
bchcmw@nus.edu.sg
Synthetic biology aims to engineer genetically modified biological systems that perform novel
functions that do not exist in nature, with reusable, standard interchangeable biological parts.
The use of these standard biological parts enables the exploitation of common engineering
principles such as standardization, decoupling, and abstraction for synthetic biology. With
this engineering framework in place, synthetic biology has the potential to make the
construction of novel biological systems a predictable, reliable, systematic process. While the
development of most synthetic biological systems remains largely ad hoc, recent efforts to
implement an engineering framework in synthetic biology have provided long-awaited
evidences that engineering principles can facilitate the construction of novel biological
systems. Synthetic biology has so far demonstrated that its framework can be applied to a
wide range of areas such as energy, environment, and health care. In this talk, our recent
efforts to develop synthetic microbes with programmable behaviors will be presented. In
particular, an emphasis will be placed on our recent development of auto-regulatory genetic
circuits for microbial chemical production and therapeutic applications.
Matthew Chang is Associate Professor at the National University of
Singapore. His research interests lie in synthetic biology of microbial
systems with particular emphasis on development of synthetic
microbes that perform programmable functions for engineering
applications. He pioneered the development of synthetic microbes that
show novel programmable therapeutic behaviors, which received
international recognition. He has been honored with the Scientific and Technological
Achievement Award from U.S. Environmental Protection Agency.
- 33 -
“MicroAntibodes”: Generation of Molecular-targeting Peptides by Directed
Evolution in Conformationally Constrained Peptide libraries
Ikuo Fujii
Department of Biological Science, Graduate School of Science, Osaka Prefecture University
fujii@b.s.osakafu-u.ac.jp
Antibodies are indisputably the most successful reagents in molecular-targeting therapy.
However, use of antibodies has been limited due to the biophysical properties and the cost to
manufacture. To enable new applications where antibodies show some limitations, we have
developed an alternative-binding molecule with non-immunoglobulin domain. The molecule is
a helix-loop-helix peptide, which is stable against enzyme degradations in vivo and is too
small to be non-immunogenic. Here, we introduce the molecular-targeting peptides termed
“microantibodes” that show antibody-like functions, high affinity and high specificity for the
targeted proteins. Since the helix-loop-helix peptide folds by virtue of the interactions
between the amino acid residues positioned inside the molecule, the outside solventexposed residues are possible to be mutated with a variety of amino acids to give a library of
the helix-loop-helix peptides. We constructed a phage-displayed library of the peptides and
screened the library against granulocyte-colony stimulating factor
(G-CSF) receptor. Finally, the screened peptide was cyclized by a
thioether linkage into the N- and C-termini. The cyclic peptide
showed a strong binding affinity (Kd of 4 nM) to the receptor and a
long half-life (>2 weeks) in mouse sera, proving an enzymeresistant property. Furthermore, immunization of the peptide to
mice showed no induction of the antibody titer (non-immunogenic).
We have applied our peptide libraries for VEGF, hIgG/Fc,
interleukins, and kinases to obtain their molecular-targeting
peptides “microAntibodies”. This semi-rational strategy, which
combines phage-displayed libraries with de novo designed
peptides, provides a new way to generate structured functional
peptides for useful tools in the field of chemical biology as well as
altanatives to antibody medicines.
Ikuo Fujii earned his Ph.D. on pharmaceutical science under the
direction of Prof. K. Kanematsu at Kyushu University (Fukuoka) in 1986,
and then he was an assistant professor at Kyushu University. He was a
postdoctoral associate in the laboratory of Prof. E. T. Kaiser at the
Rockefeller University from 1988-1989. He moved to The Scripps
Research Institute to work with Prof. R. A. Lerner in 1989. In 1991, he
was appointed to a senior research scientist to direct the antibody
engineering group in Protein Engineering Research Institute. In 2003,
he was appointed to a professor of department of biological science in
Osaka Prefecture University. At present, he is the dean of school of science. In 2013, he
received the 5th Monodzukuri Nippon Grand Award.
- 34 -
Pseudo-Natural Product Synthesis and Drug Applications
Hiroaki Suga
Department of Chemistry, Graduate School of Science, The University of Tokyo
113-0033 Tokyo, Japan
hsuga@chem.s.u-tokyo.ac.jp
The genetic code is the law of translation, where genetic information encoded in RNA is
translated to amino acid sequence. The code consists of tri-nucleotides, so-called codons,
assigning to particular amino acids. In cells or in ordinary cell-free translation systems
originating from prokaryotes, the usage of amino acids is generally restricted to 20
proteinogenic (standard) kinds, and thus the expressed peptides are composed of only such
building blocks. To overcome this limitation, we recently devised a new means to reprogram
the genetic code, which allows us to express non-standard peptides containing multiple nonproteinogenic amino acids in vitro. This lecture will describe the development in the genetic
code reprogramming technology that enables us to express natural product-inspired nonstandard peptides and pseudo-natural products. The technology involves (1) efficient
macrocyclization of peptides, (2) incorporation of non-standard amino acids, such as Nmethyl amino acids, and (3) reliable synthesis of libraries with the complexity of more than a
trillion members. When the technology is coupled with an in vitro display system, referred to
as RaPID (Random non-standard Peptide Integrated Discovery) system, the libraries of
natural product-inspired macrocycles with a variety ring sizes and building blocks can be
screened (selected) against various drug targets inexpensively, less laboriously, and very
rapidly. Moreover, the RaPID system has further evolved to a new system for the discovery
of pseudo-natural products. This lecture will discuss the most recent development of their
technology and therapeutic applications
References
1. Y. Goto, Y. Ito, Y. Kato, S. Tsunoda, H. Suga* “One-pot synthesis of azoline-containing
peptides in a cell-free translation system integrated with a posttranslational cyclodehydratase”
Chemistry & Biology, in press (2014).
2. Y. Tanaka, C.J. Hipolito, A.D. Maturana, K. Ito, T. Kuroda, T. Higuchi. T. Katoh, H.E. Kato, M.
Hattori M, K. Kumazaki, T. Tsukazaki, R. Ishitani, H. Suga*, O. Nureki “Structural basis for the
drug extrusion mechanism by a MATE multidrug transporter” Nature 496, 247-51 (2013).
3. Y. Yamagishi, I. Shoji, S. Miyagawa, T. Kawakami, T. Katoh, Y. Goto, H. Suga* "Natural
product-like macrocyclic N-methyl-peptide inhibitors against a ubiquitin ligase uncovered from
a ribosome-expressed de novo library" Chemistry&Biology 18, 1562-1570 (2011).
4. Y. Goto, T. Katoh, H. Suga “Flexizymes for genetic code reprogramming” Nature Protocols 6,
779-790 (2011)
Hiroaki Suga is a Professor of the Department of Chemistry, Graduate School
of Science in the University of Tokyo. He was born in Okayama City, Japan in
1963. He received his Bachelor of Engineering (1986) and Master of
Engineering (1989) from Okayama University, and Ph. D. in Chemistry (1994)
from the Massachusetts Institute of Technology. After three years of postdoctoral work in Massachusetts General Hospital, he was appointed as a
tenure-track Assistant Professor in the Department of Chemistry in the State
University of New York at Buffalo (1997) and promoted to the tenured Associate
Professor (2002). In 2003, he moved to the Research Center for Advanced
Science and Technology in the University of Tokyo as an Associate Professor,
and soon after he was promoted to Full Professor. In 2010, he changed his affiliation to the
Department of Chemistry, Graduate School of Science. His research interests are in the field of
bioorganic chemistry, chemical biology and biotechnology related to RNA, translation, and peptides.
He is also a founder of PeptiDream Inc. Tokyo, a publicly traded company, which has many
partnerships with pharmaceutical companies in worldwide.
- 35 -
Single organic molecule based cancer specific DDS
Byungkwon Yoon, Bomi Kwon, So Yeon Park, Soo Jin Uhm, Se Young Koo, Won
Young Kim, Kyoung Sunwoo, Jong Seung Kim*
Department of Chemistry, Korea University, Seoul, 136-701, Korea
Jongskim@korea.ac.kr
We present the design, synthesis of a new type
of targetable chemodosimetric fluorescent
sensor for imaging hepatic thiols as well as for
the drug delivery system. The in vivo and in vitro
experimental results revealed that it selectively
enters HepG2 cells as well as liver through
ASGP-R-mediated endocytosis, and shows a
fluorescence change. Thus, it is promising for
the detection/imaging of hepatic thiol levels in
vivo and for the selective delivery of anticancer
agent to the target cancer. The synthesis,
spectroscopic characterization, and biological
evaluation of a RGD peptide appended
naphthalimide pro-CPT (1) are presented. Upon
the disulfide bond cleavage reaction with free thiols, 1 displays a red-shifted fluorescence at
535 nm and releases the CPT drug concomitantly in an aqueous solution at pH 7.4. Confocal
microscopic experiments reveal that 1 is preferentially taken up by U87 cells over C6 cells
and delivers the CPT drug through a RGD-dependent endocytosis. This new DDS system
could provide a potential guideline for the construction of new therapeutic drug delivery and
its monitoring system.
References
1. Lee, M. H.; Han, J. H.; Lee, J. H.; Park, J. Y.; Jung, G. S.; Kumar, R.; Kang, C.; Kim, J. S. ACIE
6206, 52, 2013.
2. Maiti, S.; Park, N.; Han, J. H.; Jeon, H. M.; Lee, J. H.; Bhuniya, S.; Kang, C.; Kim, J. S. JACS 4567,
135, 2013.
3. Yang, Z.; He, Y.; Lee, J.-H.; Park, N.; Suh, M.; Chae, W.-S.; Cao, J.; Peng, X.; Jung, H. S.; Kang,
C.; Kim, J. S. JACS 9181, 135, 2013.
4. Yang, Z.; Lee, J. H.; Jeon, H. M.; Han, J. H.; Park, N.; He, Y.; Lee, H.; Kang, C.; Hong, K. S.; Kim,
J. S. JACS 11657, 135, 2013.
5. Lee, M. H.; Jeon, H. M.; Han, J. H.; Park, N.; Kang, C.; Sessler, J. L.; Kim, J. S. JACS 8430, 136,
2014.
6. Bhuniya, S.; Maiti, S.; Kim, E.-J.; Lee, H.; Sessler, J. L.; Hong, K. S.; Kim, J. S. ACIE 4469, 53,
2014.
7. Lee, M. H.; Park, N.; Yi, C.; Han, J. H.; Hong, J.; Kim, K. P.; Kang, D. H.; Sessler, J. L.; Kang, C.;
Kim, J. S. JACS, 2014
Jong Seung Kim received Ph. D. from Department of Chemistry and
Biochemistry at Texas Tech University. After one-year postdoctoral
fellowship at University of Houston, he joined the faculty at Konyang
University in 1994 and transferred to Dankook University in 2003. In
2007, he then moved to Department of Chemistry at Korea University in
Seoul as a professor. His research records over 300 scientific
publications and 50 domestic and international patents.
- 36 -
Chemical and Enzymatic Syntheses of Carbohydrate Oligomers
Chun-Cheng Lin1,2
1
Department of Chemistry, National Tsing Hua University, Hsinchu, Taiwan
2
Institute of Genomic Research Center, Academia Sinica, Taipei, Taiwan
cclin66@mx.nthu.edu.tw
Both chemical and enzymatic methods are the methods used in the syntheses of
complicated nature products. Chemical methods give variable pathways to synthesize
derivatives while enzymatic approaches show straightforward synthesis of specific targets. In
this talk, the uses of chemical and enzymatic methods, respectively, in the synthesis of oligosialic acid and oligo-LacNAc will be discussed.
Sialic acid is the most common non-reducing sugar of glycoproteins and glycolipids. Thus,
they act either as masks or recognition sites for the ligand-receptor and cell-cell recognitions
in many important biological events. Recently, we developed a new approach for the
synthesis of S-linked α(29)-oligo-sialic acids using an asymmetric tert-butyl disulfide
linkage as an anomeric thiol protecting group and S-alkylation of anomeric thiol nucleophile.
This strategy was further extended to synthesize S-linked α(28)-oligo-sialic acids and
alternative α(28)/α(29)-oligo-sialic acids.
Poly-N-acetyllactosamine [poly-LacNAc, (3Gal 1-4GlcNAc 1-)n] is a linear carbohydrate
polymer composed by repeating N-acetylglucosamine (GlcNAc) and galactose (Gal) residues.
These polysaccharides are found either on N-/O-linked glycoproteins or glycolipids and
involve in diverse cellular functions such as differentiation, apoptosis, and metastasis. To
efficiently and quickly produce oligo-LacNAc, we used the recombinant bacterial enzymes,
-1,3-N-acetylglucosaminyltransferase of Helicobacter pylori (HpGnT) and  -1,4galactosyltransferase of Neisseria meningitides (NmGalT), from E. coli. Defined lengths of
oligo-LacNAcs were synthesized by using the expressed transferases in the presence of
sugar donors, uridine 5‟-diphosphate galactose (UDP-Gal) and uridine 5‟-diphosphate Nacetylglucos- amine (UDP-GlcNAc). To reduce the cost of UDP-Gal and UDP-GlcNAc used
in the oligo-LacNAc synthesis, we set up an enzymatic system for one-pot synthesis of NDPsugars by using a wild-type bacterial thymidylyltransferase (RmlA). In combination with the
use of corresponding kinases, UDP-GlcNAc and UDP-Gal were obtained from cheap starting
materials, GlcNAc and Gal. Compare to previous report, we successfully achieve the
synthesis of oligo-LacNAc with a more economical way. The oligo-LacNAcs were further
modified by using sialyl transfereases and fucosyltransferase.
Chun-Cheng Lin (林俊成), National Tsing-Hua University (B 1987),
National Taiwan University (M, 1989), The Scripps Research Institute
(Ph. D. 1997); Postdoctoral Fellow, The Scripps Research Institute
(1997); Assistant Research Fellow (1998) and Associate Research
Fellow (2003), Academia Sinica; Associate Prof. (2006) and Prof. (2007)
National Tsing Hua University. Research field: carbohydrate chemistry,
enzymatic synthesis, bio-nanotechnology, protein conjugation and
immobilization.
- 37 -
How we can improve affinity of antibodies for the targets
Kouhei Tsumoto1,2,3
1
School of Engineering, 2Institute of Medical Science, and 3Open Innovation Centre for Drug
Discovery, The University of Tokyo, Japan
tsumoto@bioeng.t.u-tokyo.ac.jp
Specific recognition of ligands by proteins is a fundamental biological phenomenon, and
interaction between antigen and antibody in the immune system is a typical example. Recent
advances in physical biochemistry have enabled us to describe what factors dominated the
specificity and affinity of protein interactions. We have focused on several antigen-antibody
interactions, including those specific for model antigens, e.g. hen lysozyme, toxin, and EGFR,
and dissected the interactions from physicochemical viewpoints. Our conclusions could be
summarized as follows. 1. Specificity is dominated by only a couple of residues, named hotspot; 2. Hydrophobic interaction and/or a couple of hydrogen bonds are created by hot-spots,
and cooperative binding of other paratope residues is induced via hot-spot interactions; 3.
Other paratope residues are tolerant to site-specific mutation, and make incremental
contributions to the interaction; 4. Variable domain interactions work as a cushion for finetuning of the paratope interface; 5. Interfacial water molecules make enthalpic contribution to
the interaction, which complements the imperfect interfaces of antigen and antibody. 6. In
principle, enthalpy change dominates the high affinity of antigen-antibody interaction, e.g.
affinity maturation of the antibody. On the basis of these conclusions, we could propose one
strategic scheme on improvement of antibody affinity for targets. These conclusions could be
applied not only to improve the specificity and affinity of an antibody, but also to screen
and/or design of small molecules, which can control specific biomolecular interactions.
Kouhei Tsumoto received his BS degree in Biological Chemistry from
the University of Tokyo in 1991, and obtained PhD degree in Biological
Chemistry from the University of Tokyo in 1997. He began an academic
career at Tohoku University in Sendai, Japan (1995), and had been
involved in the antibody engineering project (1995-2005). In 2005, he
was promoted to Associate Professor of the Department of Medical
Genome Sciences, Graduate School of Frontier Sciences, the University
of Tokyo, and in 2010, he was promoted to Professor of Medical
Proteomics Laboratory, Institute of Medical Science, the University of Tokyo. Currently, he is
a full professor of bioengineering, School of Engineering, the University of Tokyo. In 2012,
the Japanese Society for the Promotion of Science (JSPS) PRIZE was awarded.
- 38 -
Intrinsic Small Molecule Fluorescent Imaging Probes towards Localized
Identification of Drug Resistant Bacterial Strains
Qing SHAO, Fang LIU, JunXin AU, Bengang XING*
Division of Chemistry and Biological Chemistry, School of Physical and Mathematical Sciences
Nanyang Technological University, Singapore
Bengang@ntu.edu.sg
So far, increased resistance of bacterial infections to antibiotic treatment has been
extensively documented and has become a generally recognized problem for clinicians
worldwide, in both hospital and community settings. Such fast growing prevalence of
antibiotic resistance may thus call for rapid and effective identification drug resistant
pathogens and systematic understanding of the biological basis towards the mode of
antibiotic binding, and the mechanism or the related genes involved in conferring bacterial
drug resistance. In general, fluorescent imaging techniques enable rapid, direct and sensitive
visualization of biological events at single cell or molecule resolution because of their high
sensitivity, relative safety, and easily handling, and therefore have become robust and
reliable tools in monitoring subcellular protein dynamics and analysis of pathogen–host
interactions. Among the different strategies for bacterial recognition, the surface affinity and
the chemical reaction-directed fluorescent labeling of resistance-associated bacterial proteins
in the context of a complicated environment offer great opportunity for the in-depth
understanding of the biological basis conferring drug resistance, and importantly, for the
facilitating of new discovery and development of effective diagnostic approaches in vitro and
in vivo. In our group, a series of simple and specific fluorescent/bioluminescent probes have
been devised and prepared to real-time visualize bacterial pathogens (Scheme 1), especially
for those with potent drug resistance through the bacterial surface ligand recognition and the
special enzyme interactions.
Scheme 1
Bengang XING received his Ph.D. in Chemistry from Nanjing University,
China, in 2000. He did his postdoctoral work with Prof. Bing Xu at
HKUST, and Prof. Jianghong Rao at Molecular Imaging Program at
University of California, Los Angles and Stanford. In 2006, he was
appointed Assistant Professor at Nanyang Technological University and
later promoted to Associated Professor in 2011. His current research
interests: Fluorescent Imaging, Chemical Biology and Nano-Medicine.
Home page: http://www.ntu.edu.sg/home/bengang/
- 39 -
Biosynthesis of Streptolidine Involved Two Unexpected Intermediates
Produced by a Dihydroxylase and a Cyclase through Unusual Mechanisms
Tsung-Lin Li
Genomics Research Center, Academia Sinica, Taiwan
tlli@gate.sinica.edu.tw
Streptothricin-F (STT-F), one of the early-discovered antibiotics, consists of three
components, a β-lysine homopolymer, an aminosugar D-gulosamine, and an unusual bicyclic
streptolidine. The biosynthesis of streptolidine is a long-lasting but unresolved puzzle. Here,
a combination of genetic/biochemical/structural approaches was used to unravel this problem.
The STT gene cluster was first sequenced from a Streptomyces variant BCRC 12163,
wherein two gene products OrfP and OrfR were characterized in vitro to be a dihydroxylase
and a cyclase, respectively. Thirteen high resolution crystal structures for both enzymes in
different reaction intermediate states were snapshotted to help elucidate their catalytic
mechanisms. OrfP catalyzes an FeII-dependent double hydroxylation reaction converting LArg into (3R,4R)-(OH)2-L-Arg via (3S)-OH-L-Arg, while OrfR catalyzes an unusual PLPdependent elimination/addition reaction cyclizing (3R,4R)-(OH)2-L-Arg to the six-membered
(4R)-OH capreomycidine. The biosynthetic mystery finally comes to light as the latter product
was incorporation into STT-F by a feeding experiment.
(n=1-7
Reference
C. Y. Chang, et al., Angew Chem Int Edit, 2014, 53, 1943-1948.
Tsung-Lin Li received his Ph.D. in Chemistry at the University of
Cambridge, UK, for work on natural products chemistry under the
supervision of the late Dr. Jonathan B. Spencer. He developed an
interest in the biosynthesis of glycopeptide antibiotics during his
postdoctoral training with Drs. Jonathan B. Spencer and Peter Leadlay in
both departments of Chemistry and Biochemistry at the same university.
After joining the faculty at National Taiwan Ocean University in 2004 he
initiated his current studies on the biosynthesis of selected natural
products. He further expanded his research after moving to the Genomics Research Center,
Academia Sinica, in 2007 by taking advantage of X-ray crystallography and synthetic biology
in a way to understand/exploit novel enzyme mechanisms that architect unique chemical
scaffolds in clinically important natural products for new drug discovery/development.
- 40 -
Small Molecule Tools for Cell Therapy
Motonari Uesugi1
1
Institutes for Integrated Cell-Material Sciences (WPI-iCeMS)
and for Chemical Research, Kyoto University, Japan
uesugi@scl.kyoto-u.ac.jp
In human history, bioactive small molecules have had three primary uses: as medicines,
agrochemicals, and biological tools. Among them, what our laboratory has done in the past
was the discovery and use of biological tools. In addition to tool discovery, our laboratory has
recently become interested in exploring another application of small molecules: small
molecule tools for cell therapy. Although small molecule drugs will continue to be important,
cell therapy will be a powerful approach to curing difficult diseases that small molecule drugs
are unable to handle. However, there are a number of potential problems in bringing cell
therapy technologies to the clinic, including high cost, potential contamination, low stability,
and tumorigenesis. Stable, completely defined small molecules, which are usually amenable
to cost-effective mass production, may be able to help the clinical application of cell therapy.
This presentation provides a quick overview of the recent results we obtained regarding
several unique molecules. These molecules were originally discovered by phenotypic cellbased screenings of our in-house chemical libraries. Molecular understanding of their
mechanisms of actions led to the design of small molecule tools that can be used both for
basic cell biology research and for cell therapy applications.
References
1. Minami, I., et al., Cell Reports, 2, 1448-1460 (2012).
2. Takemoto, N., et al., J. Am. Chem. Soc. 135, 11032-11039 (2013).
3. Sakano, D., et al., Nature Chem. Biol., 10, 141-148 (2014).
4. Hirata, N., et al., Cell Reports, 6, 1165-1174 (2014).
5. Kuo, T.F., et al., J. Am. Chem. Soc. 136, 9798-9801 (2014).
6. Frisco-Cabanos, H.L., et al., Angew. Chem. Int. Ed., in press (2014).
Motonari Uesugi is Professor of The Institute for Integrated Cell-Material
Sciences and Institute for Chemical Research, Kyoto University. After
completing postdoctoral training in Harvard Chemistry Department, Dr.
Uesugi started his independent career in Baylor College of Medicine,
Houston, where he has established an interdisciplinary laboratory in the
area of chemical biology. He was tenured in Baylor in 2005, and moved
to Kyoto University as a full professor in 2005. He is a recipient of Gold
Medal Award, Tokyo TechnoForum 21 (2006), The Pharmaceutical
Society of Japan Award for Divisional Scientific Promotions (2011) and
German Innovation Award Gottfried Wagener Prize (2011). Dr. Uesugi and his co-workers
aim to gain a fundamental understanding of biological events through the study of small
molecules. He provides the first edX course from Japan, “The Chemistry of Life,” to create a
new educational path for millions of learners worldwide.
- 41 -
. Kinome profiling of 14-membered resorcylic acid lactones and
identification of drug binding sites of drugable kinases
Taebo Sim1,2
1
Chemical Kinomics Research Center, Korea Institute of Science and Technology, Republic of Korea
2
KU-KIST Graduate School of Converging Science and Technology, Republic of Korea
tbsim@kist.re.kr / tbsim@korea.ac.kr
Naturally occurring 14-membered resorcylic acid lactones (RALs) such as hypothemycin
turned out to be covalent kinase inhibitors. In an endeavour of enriching our small molecule
kinase inhibitor discovery programs we have explored 14-membered resorcylic acid lactones
as covalent kinase inhibitors. Kinome profiling experiments for 14-membered resorcylic acid
lactones synthesized in our group have been performed to identify new molecular targets
and unique drug binding sites of drugable kinases.
Taebo Sim received his Ph.D. in Chemistry from Sogang Univ., Korea,
in 1996. He did his postdoctoral research with the late Prof. Henry
Rapoport at UC Berkeley. He commenced drug discovery research at
CKD institute in 1999 and joined the Genomics Institute of the Novartis
Research Foundation (GNF) at San Diego in 2002 and focused on
kinase drug discovery as a principal investigator. In 2006, he moved to
Dana-Farber Cancer Institute/Harvard Medical School to set-up a
medicinal chemistry lab. He joined Korea Institute of Science and
Technology (KIST) in 2007 and became the head of Chemical Kinomics Research Center at
KIST in 2012 and was appointed a professor at KU-KIST Graduate School of Converging
Science and Technology in 2012.
- 42 -
A Chemical Strategy for
Traceless Labeling and Immobilization of Glycoproteins
Po-Chiao Lin
Department of Chemistry, National Sun Yat-sen University, Taiwan
pclin@mail.nsysu.edu.tw
Glycoproteins are involved in many important biological processes such as cell development,
immunology, and cancer biology; however, only a few glycan-based targeting strategies are
currently available and most of them resulted in the alteration of native glycan structures. A
traceless strategy in labeling of glycoproteins with synthetic boronic acid (BA)-tosyl probes
was successfully developed. Due to the reversible formation of boronate ring, a competition
reaction can recover the native glycan of the tagged glycoprotein, conserving its biological
significance. To extend the concept to a chip-based study of glycoprotein-protein interactions,
a BA-tosyl–functionalized glass slide was used to fabricate glycoprotein microarrays with
highly conserved glycans. By interacting with various lectins (carbohydrate-binding proteins),
such as Concanavalin A (Con A) and wheat germ agglutinin (WGA), the types of
carbohydrates could be systematically monitored. It is believed that the newly developed
method will accelerate the understanding of glycoproteins. Recently, the labeling efficiency
and specificity of glycoproteins was found to be considerably improved by modifying the
structures of the benzenesulfonyl reactive center.
References
1. Y.-L. Yang, Y.-P. Lee, Y.-L. Yang, P.-C. Lin* ACS Chem. Biol. 2014, 9, 390-397.
2. Y.-L. Yang, B.r Rajagopal, C.-F. Liang, C.-C. Chen, H.-P. Lai, C.-H. Chou, Y.-P. Lee,
Y.-L. Yang, J.-W. Zeng, C.-L. Ou, P.-C. Lin,* Tetrahedron 2013, 69, 2640-2646.
Po-Chiao Lin (1979, PingTung, Taiwan) received his PhD at Taiwan
International Graduate Program in 2008 with Prof. Chun-Cheng Lin. In
2010, He completed a post-doctoral appointment with Prof. Herbert
Waldmann at Max-Planck-Institute for Molecular Physiology (Dortmund,
Germany) and was appointed as Assistant Professor of Organic
Chemistry at National Sun Yat-sen University. His research interests are
in organic synthesis, protein engineering and protein microarray.
- 43 -
Glyco-based interplay between Helicobacter pylori and gastric epithelium
Chun-Hung Hans Lin
Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan
Institute of Biochemical Science, College of Life Science, National Taiwan UNiversity
chunhung@gate.sinica.edu.tw
We herein report the two interplays of Helicobacter pylori with gastric epithelium. First,
using fluorescently labeled fucose-containing glycoconjugates, we provide evidence
observing both the uptake of L-fucose from gastric cancer cells to H. pylori and that human
-L-fucosidase 2 (FUCA2) is secreted only when host cells were infected with H.
pylori. FUCA2 was found to be essential for H. pylori adhesion, in particular to the gastric
cancer- and duodenal ulcer-specific strains. FUCA2 was also shown to significantly enhance
the expression of Lewis x antigen in H. pylori, which is critical for bacterial cell adhesion in
the pathogenesis and defense strategy to escape host surveillance. In addition, potent
fucosidase inhibitors were utilized to effectively abolish the bacterial infection, supporting the
idea that FUCA2 could be a potential target for therapeutic intervention. We will also describe
the development of fucosidase inhibitors, as well as the corresponding x-ray structures to
understand the binding interactions at molecular basis.
The other interplay is relevant to cholesteryl glucosides. It is known that H. pylori cannot
synthesize cholesterol. The pathogen not only takes up host cholesterol, but also modifies it
into different forms of cholesteryl glucosides. We will report the development of a specific
metabolite-labeling method for the purpose of rapid identification, quantitative analysis and
structural characterization.
References
1. T.-W. Liu, H.-H. Huang, C.-W. Ho, S.-M. Chang, S. S. Popat, M.-S. Wu, Y.-J. Chen, C.-H.
Lin. Proc Natl Acad Sci USA 2009, 106, 14581-14586.
2. H.-J. Wu, C.-W. Ho, T.-P. Ko, S. S. Popat, C.-H. Lin, A. H.-J. Wang Angew Chem Int Ed.
2010, 49, 337-340.
3. Z. Tu, Y.-N. Lin, C.-H. Lin Chem Soc Rev 2013, 42, 4459-75.
Chun-Hung Hans Lin received his Ph.D. in Chemistry at the Scripps
Research Institute, La Jolla, USA, in 1995 for work on enzymatic
synthesis of carbohydrates under the supervision of Dr. Chi-Huey Wong.
He studied the enzyme reaction mechanism under the instruction of Dr.
Christopher T. Walsh at Harvard Medical School in 1995-97. He started
his independent career at the Institute of Biological Chemistry,
Academia Sinica as Assistant Res Fellow (1998-2002), Associate Res
Fellow (2002-07), and Res Fellow (since 2007). His major interest is to
develop selective inhibitors/probes for several proteins, including
fucosidase, fucosyltransferase, glutathione-conjugated proteins. The
development led to the identified interplays between Helicobacter pylori
and gastric epithelium. He received several awards and honors, including Young Affiliate of
TWAS (2008-12), The 2010 Young Scholar Award of Tien-De Li Biomedical Foundation,
Investigator Award of Academia Sinica (2011), NSC Outstanding Research Award in 2013.
- 44 -
Glycosylation of flavonoids and polyketides using flexible glycosyltransferase
Jae Kyung Sohng
Department of BT-convergent Pharmaceutical Engineering, Sun Moon University, 70 Sunmoonro-221,
Tangjeonmyun, Asansi, Chungnam 336-708, Republic of Korea
sohng@sunmoon.ac.kr
A UDP-glycosyltransferase, YjiC from Bacillus licheniformis DSM13 was exploited for the
glycosylation of a number of small molecules. The in-vitro glycosyltransferase assay using
YjiC lead to the production of multiple glucosides of polyhydroxyl groups containing
aromatic/phenolic as well as aliphatic small molecules exhibiting wide substrate flexibility of
the enzyme. We tested the activity of the enzyme with different classes of flavonoids
(flavonoid, isoflavonoids, stilbenes, xanthonoids, benzoates) and polyketides (anthraquinone
and macrolide). Moreover, we have found the wide donor substrate flexibility of YjiC with
different NDP-sugars transferring a range of modified sugars including L- and D-sugars to
the selected acceptors which guide to the production of a number of novel derivatives of
natural products. But, it is found that YjiC could transfer only at limited positions of the
polyhydroxylated compounds with diverse NDP-sugar donors. We further successfully
applied YjiC for the in-vivo biotransformation of different flavonoids using engineered E. coli
in large scale fermentor to produce glucosylated derivatives. In conclusion, it is found that
YjiC has wide donor as well as acceptor substrates flexibility with high catalytic activity. The
further exploitation of such glycosyltransferases could help to efficient production of
glycosylated derivatives of natural products which might have potential therapeutic as well as
cosmetic applications.
Jae Kyung Sohng
Email: sohng@sunmoon.ac.kr
Lab: +82-41-530-2246
▪ Education
B.S., Yonsei University, Dept of Chemistry, 1979-1984
M.S., Yonsei University, Dept of Chemistry, 1984-1986
Ph.D, Brown University, Dept of Chemistry, 1986-1991
▪ Carrier
July 1991 - Feb 1994: University of Washington, (Post-doc)
March 1994 – present: SunMoon University (Professor)
March 2010 – Feb. 2012: College of Health Science, SunMoon University (Dean)
January 2010 – present: Asian Communications of Glycobiology and Glycotechnology (Steering
Committee)
▪ Recent Publication
1. “The 7th Japan-Korea chemical biology symposium: chemical biology of natural bioactive
molecules.” ACS Chem Biol. 2014, 16, 1070-4.
2. “Methylation and subsequent glycosylation of 7,8-dihydroxyflavone” J Biotechnol. 2014,184,128-137.
3. “Assessing acceptor substrate promiscuity of YjiC-mediated glycosylation toward flavonoids.”
Carbohydr Res. 2014, 1, 393:26-31.
4. “Efficient enzymatic systems for synthesis of novel α-mangostin glycosides exhibiting antibacterial
activity against Gram-positive bacteria.” Appl Microbiol Biotechnol. 2014, Jul 20.
5. “Recent advances in biochemistry and biotechnological synthesis of avermectins and their
derivatives” Appl Microbiol Biotechnol. 2014, Jul 20.
- 45 -
Microbial glycoconjugates and lipidconjugates as immunomodulators;
Chemical synthesis and biological functions
Yukari Fujimoto
Department of Chemistry, Faculty of Science and Technology, Keio University
fujimotoy@chem.keio.ac.jp
Many of microbial glycoconjugates and lipidconjugates from the surface of microbe cells are
known as immunomodulators. We have been interested in the precise chemical syntheses
and the functional analysis of these compounds. One of the major components is bacterial
cell wall peptidoglycan (PGN), which has common structural motives including glycan chains
of alternating GlcNAc-MurNAc with L-Ala-D-Glu as a unit structure. Presumably because of
the structural commonality, macrocellular organisms recognize the PGN with various proteins
of their immune system, and bacteria obviously have many kinds of proteins to recognize the
PGN, as their biosynthetic target or for anchoring to the cell wall. We have synthesized
various PGN fragment structures to make the compound library for applying to the functional
analysis of the molecule. One of the application is the array of PGN fragment structures; It
enabled to analyze the recognition site of PGN by the various proteins.
References:
1. a) Fujimoto, Y., et al., Nat. Prod. Rep. 2012, 29, 568;
b) Fujimoto, Y., et al., J. Endotoxin Research 2007, 13, 189.
2. Wang, N., et al., ChemBioChem, 2013, 14, 482.
3. Mesnage, S., et al., Nat. Commun. 2014, 5:4269.
Yukari Fujimoto is a Professor of Department of Chemistry, Faculty of
Science and Technology, Keio University. She received her B.S. from
Osaka University in 1989. After industrial experience at Sumitomo
Chemical Co., Ltd. and post-graduate studies at Columbia University,
USA, she received her Ph. D. from Osaka University in 2002. After
postdoctoral studies at Nagoya University, she joined the faculty of
Osaka University as an Assistant Professor in 2003, where she
promoted to a Lecturer (2006) and an Associate Professor (2008). In
April, 2014, she moved to Keio University as a full professor. Her current
major interest is in the area of chemical synthesis and functional
analysis of immunostimulatory compounds including microbial glycoconjugates and
glycolipids.
- 46 -
Labeling and Visualizing Glycans with Specificity and Versatility
Xing Chen
Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking
University, Beijing, 100871, P.R. China
xingchen@pku.edu.cn
Glycosylation plays a key role in mediating molecular recognition, development, and cell
signaling. Our research group works on developing chemical tools to label, visualize, and
modulate glycosylation. Here, we present the development of a liposome-based strategy for
cell-selective metabolic labeling of glycans and a FRET-based method for protein-specific
imaging of cell-surface glycans.[1,2] Furthermore, we design and synthesize bifunctional
unnatural sugars that introduce simultaneously two functional groups into cellular glycans.[3]
The bifunctional sugar analogs are valuable tools for profiling glycan-protein interactions.
Finally, we are interested in developing new imaging modalities for glycan visualization. A
recently developed bioorthogoal Raman imaging technique will be discussed.[4,5]
References:
1. Lin, W.; Du, Y.; Zhu, Y.; Chen, X. J. Am. Chem. Soc. 2014, 136, 679-687.
2. Xie, R.; Hong, S.; Feng, L.; Rong, J.; Chen, X. J. Am. Chem. Soc. 2012, 134, 99149917.
3. Feng, L.; Hong, S.; Rong, J.; You, Q.; Dai, P.; Huang, R.; Tan, Y.; Hong, W.; Xie, C.;
Zhao, J.; Chen, X. J. Am. Chem. Soc. 2013,135, 9244-9247.
4. Lin, L.; Tian, X.; Hong, S.; Dai, P.; You, Q.; Wang, R.; Feng, L.; Xie, C.; Tian, Z.;
Chen, X. Angew. Chem. Int. Ed. 2013, 52, 7266-7271.
5. Hong, S.; Chen, T.; Zhu, Y.; Li, A.; Huang, Y.; Chen, X. Angew. Chem. Int. Ed. 2014,
53, 5827-5831.
Dr. Xing Chen was born in October 1980. He completed his
undergraduate degree in Chemistry from Tsinghua University in 2002.
Dr. Chen then obtained his Ph.D. in Chemistry from University of
California, Berkeley in 2007, where his research focused on chemical
biology and bionanotechnology. After completing postdoctoral work at
Harvard Medical School in the field of structural biology and
immunology, he joined the Peking University faculty in September, 2010.
Some of his awards include: Chinese Chemical Society Young Chemist
Award (2013); DuPont Young Professor Award (2013); SCOPUS Young Researcher Award
(2012); The current research interest of Dr. Chen is focused on chemical glycobiology.
- 47 -
Bio-recognition and Bio-sensing---From big to small
Bor-ran Li, Mo-Yuan Shern, and Yaw-Kuen Li*
Department of Applied Chemistry, National Chiao Tung Universty,
1001 University Rd. Hsin-Chu, Taiwan 30010
ykl@faculty.nctu.edu.tw
Biosensors have drawn much attention because of their potential to greatly improve
biomedical research, drug discovery, environmental monitoring, and diagnosis of many
diseases. A powerful bio-detection requires a sensitive sensing device and a highly specific
bio-recognition apparatus. Several techniques such as gold nano-particles (AuNPs), quartz
crystal microbalance (QCM), and, surface plasmon resonance (SPR), silicon nanowire fieldeffect transistor (SiNW-FET) and others have been extensively studied. Yet, efficient
methods for bio-conjugation on sensor chip and for leasing the non-specific interaction of
biomolecule from sensor are remained to be solved. A new approach to directly modify the
zwitterionic molecule, dimethylammonio propane-1-sulfonate (sulfobetaine or SB), on the Au
surface of QCM chip via electrodeposition is demonstrated. The zwitterion-functionalized
antifouling surface enables a decrease by 95 % of the adsorption of non-specific proteins
from fetal bovine serum (FBS, 10 %). For bio-recognition, the animal antibody is commonly
used to couple with the above devices. This conventional technique is good for big molecule
but not for small one. In this presentation, the technique for screening scFv for the effective
open sandwich recognition from phage display library will also be discussed. It is a powerful
technique for establishing a small molecule as well as a big molecule detection.
Prof. Yaw-Kuen Li received his Ph.D. in Chemistry from Tulane
University, USA, in 1991. He performed his postdoctoral research
with Prof. Paul Talalay at Johns Hopkins School of Medicine in
1991-1993 and further became a faculty member of Applied
Chemistry Department at National Chiao Tung University, Taiwan,
in 1993. He served as a chair of the department in 2004-2006 and
assigned as the project director of chemistry division of National
Science Council of Taiwan in 2011-2013. Currently, he is the team
leader of Biomolecular Probing and the dean of College of Science
of the university.
- 48 -
Fusarisetin A, A Novel Cancer Cell Migration Inhibitor from Fusarium sp.
FN080326, and It‟s Cellular Binding Protein
Jong Seog Ahn
Chemical Biology Research Center, Korea Research Institute of Bioscience and Biotechnology,
(KRIBB) Ochang, Choongbuk, Korea
jsahn@kribb.re.kr
Secondary metabolites produced by microorganisms have proven to be a valuable repository
of natural bioactive compounds and many of them have been identified as useful research
reagents and potential drug candidates. Traditionally, a lot of bioactive molecules have been
reported from Actinomyces strains, but recently the number of novel bioactive molecules
isolated from fungal strains is increasing.
In the course of screening soil fungal isolates, we discovered a novel compound named
fusarisetin A (FSA) from a soil fungus, Fusarium sp. FN080326, showing inhibitory activity on
cancer cell motility. From the chemical structure determination, we know that FSA possesses
an unprecedented carbon skeleton with a pentacyclic ring system comprising a decalinemoiety (6/6) and a tricyclic moiety (5/5/5). FSA effectively inhibited the 3D-culture multiaciniform and acini-colony of MDA-MB-231 (human breast cancer cells). And the cell
migration was encumbered by FSA in a dose-dependent manner without any significant
cytotoxicity. Moreover, FSA strongly interfered with the 10% FBS, EGF, or bFGF-induced cell
invasion in MDA-MB-231 cells. By the pull-down of cell lysate using FSA-immobilized beads,
a candidate binding protein for FSA could be identified. The gene knock out experiments
(siRNA and shRNA) and over-expression for the candidate protein in MDA-MB-231cells
showed the same phenotypic effects of FSA treatment such as inhibition of acinar
morphogenesis and tumor cell invasion. So we suggested that FSA has a potential for antimetastatic agent and the candidate protein can be explored as a new molecular target for
cancer metastasis
Reference
1. Jang JH, et al J. Am. Chem. Soc. 20111, 133: 6865-6867.
2. J. Xu, et al. Chem. Sci. 2012, 3, 3378-3386.
Jong Seog Ahn graduated from Dept. Biology of Seoul National University
(1979) and received Ph. D in Biotechnology from Korea Advanced
Institute of Science and Technology (1985). And then he started his
works as a senior research scientist at KRIBB and currently he is the
director of Chemical Biology Research Center of KRIBB and a professor
of University of Science and Technology. He also had the chances to
work in Antibiotics Lab. of RIKEN (1986-1987) and in Medical School of
Duke University (1999) as visiting scientist.
- 49 -
Poster Abstracts
- 50 -
P-01
Engineering hemolytic peptides for intracellular delivery of biomacromolecules
Misao Akishiba, Toshihide Takeuchi, Yoshimasa Kawaguchi and Shiroh Futaki
Institute for Chemical Research, Kyoto University, Japan
akishiba.misao.87m@st.kyoto-u.ac.jp
Intracellular delivery of various membrane-impermeable molecules is in great demand for
development of biopharmaceuticals. Approaches to achieve efficient cellular internalization of large
molecules should contribute to the development of antibody therapies, which have now received much
attention because of high specificity of antibodies in molecular recognition. Many approaches have
been reported up to the present time. However, few of them are efficient enough to deliver highmolecular-weight proteins into cytosol to effectively modulate cell functions.
In this study, we designed a peptide to efficiently deliver bioactive proteins into cytosol by disrupting
endosomal membrane. Amphipathic α-helical conformation is widely observed for antimicrobial
peptides which can injure plasma membrane. By modifying amphipathic α-helical sequences of natural
hemolytic peptides, we have succeeded in creating a peptide which has low cytotoxicity but has an
enough ability of endosome disruption yielding marked cytosolic diffusion of endocytosed protein.
Applicability of this peptide to cytosolic delivery of antibodies will also be discussed.
P-02
Plant-Based Polymeric Materials Design using a Sugar-diol as a Pendant Group
Hidenobu Takao1, Yuzo Ishigaki2, Keigo Aoi1
1
Graduate School of Bioagricultural Sciences, Nagoya University, Japan
2
Nagoya Municipal Industrial Research Institute, Japan
aoi@agr.nagoya-u.ac.jp
In relation to the field of chemical biology, macromolecular design of biomaterials is important to
investigate and apply the biological and chemical events. As for the biomaterials, polymeric materials
using naturally abundant carbohydrates have an advantage in an ecological viewpoint besides fully
artificial molecular design. We have studied syntheses and applications of polyesters, poly(esteramide)s, poly(ester-carbonate)s, and polycarbonates containing isosorbide, i.e., 1,4:3,6-dianhydro-Dglucitol, which has a fused tetrahydrofurane (THF) ring structure and is derived from starch as an
industrial by-product.
Nowadays isosorbide is commercially available from D-glucose by
hydrogenation, followed by dehydration.
We have already reported isosorbide-containing
1
polycarbonates can be utilized as Li ion-conductive materials . Although isosorbide is a promising diol
in condensation reactions, we designed pendant-type structure due to apply isosorbide-containing
polymers to high-performance ion-conductive materials.
In the present report, we will show synthesis and properties of novel polyacrylates having the
isosorbide unit as a pendant group. Molecular motion of the side isosorbide group should be high to
carry Li ion efficiently. The polyacrylates were synthesized by atom-transfer radical polymerization
(ATRP) of isosorbide having acryl esters. One hydroxyl group of the isosorbide is connected to the
vinyl group via ester linkage, whereas the other hydroxyl group is chemically modified to methoxy
group and several acyloxy groups. Therefore, the polyacrylates have side isosorbide groups with
various alkyl chains. Molecular weights of the obtained polymers were around 20,000 estimated by
size exclusion chromatography (SEC). Their glass transition temperatures determined by differential
scanning calorimetry (DSC) decrease systematically with increasing the alkyl chain lengths. Li ion
conductivity of the polymers was measured with Li TFSI. Relationship between the chemical structure
and activation energy will be also discussed.
1 Y. Saito, K. Hirai, H. Katayama, T. Abe, M. Yokoe, K. Aoi, M. Okada, Macromolecules, 2005, 38,
6485.
- 51 -
P-03
Fluorescent Probes for Targeted Visualization of Temperature at Organelles in single
living cells
Satoshi Arai1, Madoka Suzuki1,2, Young-Tae Chang3,4
1
WASEDA Bioscience Research Institute in Singapore (WABIOS), Singapore. 2Organization for
University Research Initiatives, Waseda University, Japan. 3Laboratory of Bioimaging Probe
Development, Singapore Bioimaging Consortium, Agency for Science, Technology and Research
(A*STAR), Singapore. 4Department of Chemistry, National University of Singapore, Singapore.
s.arai@kurenai.waseda.jp
An optical probe called fluorescent thermometer, which reports small temperature differences as a
fluorescent signal, has garnered attentions as a promising tool for intracellular thermometry. Herein we
describe the first small molecule fluorescent thermometers selectively targeting organelles responsible
for heat production. We screened the diversity oriented fluorescent library (DOFL) for temperature
sensitive dyes and also those targeting organelles. We finally selected small thermometers that
localized to endoplasmic reticulum (ER) and mitochondria as well-known organelles for heat
production, and termed them as ER and Mito thermo yellow, respectively. Unlike nanoparticle
thermometers, these novel thermometers stain target organelles evenly and successfully visualize the
intracellular temperature gradient generated by external heat source in various cell types. We further
2+
confirm the ability of ER thermo yellow in monitoring of the heat production by intracellular Ca
change in HeLa cells, validating the feasibility of physiological monitoring of the temperature change at
the subcellular level. The temperature imaging at the organelles will provide the precise view of
thermogenesis at subcellular level and shed light on the crosstalk between each organelle as well as
individual cells.
P-04
Dual-targeting delivery systems: selective cancer cell death and imaging
Kyung-Hwa Baek1, Xizhe Tian1,2 and Injae Shin1
1
National Creative Research Center for Biofunctional Molecules, Department of Chemistry, Yonsei
University, Seoul 120-749, Korea
2
Department of Chemistry, College of Sciences, Yanbian University, Yanji 133000, China
injae@yonsei.ac.kr
Although many anticancer agents have developed to treat cancer, these show many unwanted normal
cell death because of deficiency of selectivity on cancer cell. Additionally, tumor imaging agents also
exhibit low target-to-background ratios. The elegant methods that more specifically target cancer cells
need to be developed for the improvement of chemotherapeutic efficacy and diagnosis. Here we
designed, synthesized and explored the effectiveness of a dual-targeting delivery system that targets
cancer cells more selectively, in an effort aimed at improving the tumor selectivity of therapeutic and
imaging agents. The new delivery system is composed of a synthetic ligand (octreotide) of
somatostatin receptors, a dipeptide substrate for cathepsin B, and a fluorophore or an anticancer
agent. The fluorophore-conjugated delivery system was found to be applicable for specific
fluorescence imaging of cancer cells that express both somatostatin receptors and cathepsin B. In
addition, the anticancer agent containing delivery system leads to the death of cancer cells specifically.
In contrast to cancer cells, normal cells that do not produce both somatostatin receptors and cathepsin
B at high levels are unaffected by the delivery system. The new dual-targeting approach has the
capability of overcoming obstacles associated with current chemotherapeutic and imaging methods.
- 52 -
P-05
Investigating the Mechanism of Lipid A Binding to CD14 using Molecular Dynamics
Simulations
Nils A. Berglund1,2, Syma Khalid2 and, Peter J. Bond1
Bioinformatics Institute, A*STAR, Singapore,138671, Singapore.
School of Chemistry, University of Southampton, Highfield, Southampton, SO17 1BJ, UK.
nilsab@bii.a-star.edu.sg, peterjb@bii.a-star.edu.sg.
The CD14 co-receptor is specialized for recognition of bacterial lipopolysaccharide (LPS). On the
surface of macrophages and other immune cells, it transfers LPS and its bioactive component lipid A
to the MD-2 protein in complex with Toll-Like Receptor 4 (TLR4), and is hence crucial in activating the
innate immune system via the TLR4 signalling pathway. In the case of severe infections, lipid A can
cause sepsis through over-activation of the immune response, leading to multiple organ failure and
death, and has become a major target for anti-septic drugs. Unfortunately, the mechanism by which
lipid A is transferred to CD14, and the detailed mode(s) of associated binding, are unknown. In this
study we have used atomically detailed molecular dynamics simulation approaches to uncover the
mechanism by which lipid A is transferred to, interacts with and binds to a hypothesized aminoterminal pocket in CD14. We modelled the interactions and dynamics of CD14 in the presence of a
range of lipid ligands, including control fatty acid systems, and lipid A in monomeric and
aggregate/micelle forms. These simulations were run in order to observe the spontaneous ligand
binding process, and have subsequently been extended to establish the thermodynamics of ligand
recognition. Our results emphasise the dynamic nature of the amino-terminal pocket which allows it to
adapt its volume to widely varying ligand size, consistent with the broad specificity of CD14. We have
also identified a possible ligand gating mechanism consistent with available NMR data, and key sites
that may be essential for LPS/lipid A binding which may ultimately be targeted by novel anti-septic
drugs.
P-06
Enzymatic Introduction of PEG Strands onto DNA and Effects of the PEG Modification
on Cell Adhesion
Hiroto Fujita1, Kosuke Nakajima1, Yuuya Kasahara1,2, Masayasu Kuwahara1
1
Graduate School of Science and Technology, Gunma University, Japan
2
National Institute of Biomedical Innovation (NIBIO), Japan
mkuwa@gunma-u.ac.jp
We enzymatically synthesized a PEG-modified DNA aptamer that could bind to human thrombin and
analyzed its target-binding affinities using a nonequilibrium capillary electrophoresis of equilibrium
mixtures (NECEEM) method. This modified DNA aptamer had target binding activity comparable to
that of the original natural DNA aptamer. A PEG-modified DNA aptamer–thrombin complex showed a
considerably reduced protease activity as compared with that of free thrombin, but it could generate
fibrin polymers. During fibrin generation, the fluorescence polarization of a FAM-labeled thrombinbinding DNA aptamer (TBA) was monitored in the presence of thrombin. TBA was observed to be
selectively trapped within fibrin polymers during gel growth, which was catalyzed by thrombin. We
designated this phenomenon as “selective oligonucleotide entrapment in fibrin polymers” (SOEF).
Furthermore, using SOEF, we investigated multiple incorporations of amphiphilic aliphatic groups into
fibrin gels via PEG-modified TBA and its effects on cell adhesion.
1. Kasahara, Y.; Irisawa, Y.; Fujita, H.; Yahara, A.; Ozaki, H.; Obika, S.; Kuwahara, M. Anal. Chem.
2013, 85, 4961–4967.
2. Undas, A. Thromb. Haemost. 2014, 112, 32–42.
3. Wood, M. D.; MacEwan, M. R.; French, A. R.; Moore, A. M.; Hunter, D. A.; MacKinnon, S. E.;
Moran, D. W.; Borschel, G. H.; Sakiyama-Elbert, S. E. Biotechnol. Bioeng. 2010, 106, 970–979.
- 53 -
P-07
Efficient Labeling of Amino Groups by Azaelectrocyclization, Application to PET and
Fluoresecent Imaging of Biomolecules and Living Cells.
Koichi Fukase1 and Katsunori Tanaka2
1
Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan
RIKEN, Biofunctional Synthetic Chemistry Laboratory, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan
koichi@chem.sci.osaka-u.ac.jp
2
We have reported the efficient labeling of peptides, proteins, and living cells by using 6π-8
azaelectrocyclization under mild conditions (10-30 min, 24-37°C, 10 M) and applied this method for
PET and fluorescent imaging of biomolecules and living cells [1-3]. We then successfully visualized
the N-glycan-dependent bio-dynamics of the labeled glycoproteins, glycodendrimers, and living cells in
the whole-body system.
P-08
Synthetic Study of Clickable Dysiherbaine
Koichi Fukushima, Masato Oikawa
Graduate School of Nanobioscience, Yokohama City University, Japan
moikawa@yokohama-cu.ac.jp
Chemical probes are important tools to
elucidate functions of target proteins as well as
their interactions. Development of chemical
probes for ionotropic glutamate receptor
(iGluRs), that mediates majority of fast
excitatory neurotransmission in the central
nervous system (CNS), is highly challenging
due to the unique structure of the ligand
binding domain.
Dysiherbaine (DH, Fig. 1), which was isolated from Micronesian marine sponge Lendenfeldia
chondrodes in 1997 by Sakai et al, is a potent agonist selective to GluK1- and GluK2-containing
subtypes of kainate type iGluR. In the present study, we planned to synthesize clickable DH 1 (Fig. 1)
as a precursor for the chemical probe which has ethynyl group at the C6 position. While several total
syntheses of DH have been reported, de novo synthetic route was needed to be developed for the
clickable DH bearing additional quaternary carbon center at C6.
Herein we report stereoselective synthesis of the vital intermediate, which has three stereocenters,
performed in 10 steps starting from D-ribose. The key transformation includes domino aldolCannizzaro reaction followed by stereoselective aldol reaction at the ring juncture of the
bicyclo[3.3.0]octane skeleton, and stereoselective addition of ethynyl group to the lactol moiety.
- 54 -
P-09
Ligand-dependent active site closure revealed in the crystal structure of
Mycobacterium tuberculosis MenB complexed with product analogs
Haigang Song1, Hoi Pang Sung1, Yuk Sing Tse1, Ming Jiang1, 2, Zhihong Guo1
1
Department of Chemistry and State Key Laboratory of Molecular Neuroscience, The Hong Kong
University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, China. 2Current
address: State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology,
Shanghai Jiao Tong University, Shanghai 200240, China.
chguo@ust.hk
1, 4-Dihydroxy-2-naphthoyl coenzyme A (DHNA-CoA) synthase catalyzes an essential intramolecular
Claisen condensation in menaquinone biosynthesis and is an important target for development of new
antibiotics. This enzyme in Mycobacterium tuberculosis is cofactor-free and classified as a type II
DHNA-CoA synthase, which is different from type I enzymes that rely on exogenous bicarbonate for
catalysis. Its crystal structures in complex with product analogs have been determined at a high
resolution to reveal ligand-dependent structural changes, which include ordering of a 27-residue
active-site loop (aa. 107-133) and reorientation of the carboxy terminal helix (aa. 289-301) that forms
part of the active site from the opposing subunit across the trimer-trimer interface. These structural
changes result in closure of the active site to the bulk solution likely through an induced-fit mechanism,
similar to what have been observed for the type I DHNA-CoA synthases. These findings demonstrate
that the ligand-dependent conformational changes are a conserved feature of all DHNA-CoA
synthases, gleaning new insights into the catalytic mechanism of the essential tubercular enzyme.
Acknowledgements: This work was supported by GRF601413, GRF601203, and FSGRF13SC01
from the Research Grants Council of the HKSAR government. We thank SSRF (Shanghai, China) for
access to beamline BL17U1 and the beamline staff for technical support.
P-10
Study on the Target Proteins of Antitumor Macrolide Aplyronine A
Yuichiro Hirayama1, Kozo Yoneda1, Kota Yamagishi1, Kota Tsuchiya1, Takumi Chinen2,
Takeo Usui2, Eriko Sumiya3, Motonari Uesugi3, Tomohiro Suzuki4, Hirokazu Kawagishi4,
Masaki Kita1, Hideo Kigoshi1
1
Graduate School of Pure and Applied Sciences, University of Tsukuba, Japan
Graduate School of Life and Environmental Sciences, University of Tsukuba, Japan
3
Institute for Integrated Cell-Material Sciences and Institute for Chemical Research, Kyoto University, Japan
4
Graduate School of Science and Technology, Shizuoka University, Japan
hirayama@dmb.chem.tsukuba.ac.jp
2
Aplyronine A (ApA) is a marine natural product that shows potent antitumor activity. The primary target
of ApA was found to be actin. However, studies on the structure-activity relationships and X-ray
analysis of the actin–ApA complex suggested that the potent cytotoxicity of ApA was not entirely
accounted for by only interaction of actin, and its molecular targets and mechanisms of action
remained unclear. To identify the additional target molecules of ApA, photoaffinity biotin derivatives of
ApA were synthesized. By the photolabeling experiments in the living tumour cells, tubulin was
identified as a secondary target of ApA. To clarify the interactions among ApA, actin and tubulin, in
vitro assays, such as gel-permeation HPLC and surface plasmon resonance analyses, were
performed. These experiments revealed that ApA forms a 1:1:1 heterotrimeric complex with an actin
monomer and a tubulin heterodimer to potently inhibit tubulin polymerization. The detail studies of the
target identification of ApA and its unique interaction with the two cytoskeletal proteins will be
presented.
- 55 -
P-11
Proteomic Analysis on Enhanced Tolerance of Nanoencapsulated Saccharomyces
Cerevisiae against UV-C Irradiation
Daewha Hong, Hojae Lee, Taegyun Park, Hee Chul Moon, Insung Choi*
Center for Cell-Encapsulation Research, Department of Chemistry, KAIST, Daejeon 305-701, Korea
dw_hong@kaist.ac.kr
In this work, we used a cytocompatible two-step process to encapsulate Saccharomyces cerevisiae in
a highly uniform nanometric (< 100 nm) shell composed of organic poly(norepinephrine) and inorganic
silica layers. The resulting cell-in-shell structure showed the structural mimicry of bacterial endospores
and acquired enhanced tolerance against germicidal UV-C irradiation as well as lytic enzyme and
desiccation. Along with the structural transformation, mass spectrometry-based proteomic analysis
indicated that S. cerevisiae altered their metabolic states during the encapsulation process, and this
biochemical adaptation of S. cerevisiae contributed to the observed UV-C tolerance, in addition to the
shell‟s UV-C filtering property. Especially, the molecular chaperons of the HSP70 family (ECM10 and
SSA1) were found to be up-regulated only for the encapsulated S. cerevisiae upon the UV-C exposure.
This work suggests a chemical tool for cytoprotecting individual living cells and also manipulating the
cellular activities at a single-cell level.
P-12
Chemoenzymatic synthesis of cyclic sialyl Lewis X 6-sulfate and analogues
Chuen-Jiuan Huang
Institute of Biological Chemistry, Academia Sinica
h291172@gate.sinica.edu.tw
Selectin-mediated cell adhesion is essential for lymphocyte homing and leukocyte recruitment into
X
immune foci. Sialyl Lewis X (sLe ), the carbohydrate ligand of E- and P-selectins, is constitutively
X
expressed on monocytes and granulocytes. Most peripheral resting T cells do not express sLe ;
X
however, they are strongly induced to express sLe upon activation. In contrast, a subset of peripheral
resting memory T helper cells expresses another carbohydrate ligand for E- and P-selectins, namely
X
X
sialyl 6-sulfo Le . Interestingly, the selectin-binding activity of sialyl 6-sulfo Le is abrogated due to the
X
cyclization of its sialic acid moiety. Briefly, the N-acetylneuraminic acid residue of sialyl 6-sulfo Le , is
de-N-acetylated to form an amino group, followed by an amide-forming reaction between the amino
group and the C1-carboxylic acid to form the cyclized neuraminic acid. Because these reactions
modify the sialic acid that is critical to the interaction with selectins, the final product, cyclic sialyl 6X
sulfo Le , loses the selectin-binding activity, indicating an inactivating mechanism of selectin binding
activity at the cell surface during the leukocyte extravasation. Herein we report the development of an
expeditious synthesis of cyclic sialyl Lewis X 6-sulfate and analogues. Our studies intend to address
two important issues. (1) The incorporation of sulfate appeared to interfere with the enzymatic
synthesis. Several approaches were tried for satisfying solutions. (2) The synthesis of sialyl Lewis x
analogues allow us to investigate what factors (the presence of sulfate, L-fucose, etc) plays a role in
affecting the lactam formation. Several methods including HPLC and NMR have been utilized to
measure the rate of sialic acid cyclization. The results will be shown and discussed.
- 56 -
P-13
Cell-Penetrating, Dimeric a-Helical Peptides: Nanomolar Inhibitors of
HIV-1 Transcription
Sangmok Jang1, Soonsil Hyun1, Seoyeon Kim1, Seonju Lee2, Im-Soon Lee3, Masanori Baba4,
Yan Lee2,* and Jaehoon Yu*,1
1
Department of Chemistry & Education, Seoul National University, Seoul, Korea
2
Department of Chemistry, Seoul National University, Seoul, Korea
3
Department of Biological Science, Konkuk University, Seoul, Korea
4
Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan
jhoonyu@snu.ac.kr and gacn@snu.ac.kr
We constructed dimeric α-helical peptide bundles based on leucines (L) and lysines (K) for both
efficient cell penetration and inhibition of Tat-TAR interaction. Owing to their ca. 90% of helical
contents in an aqueous buffer, the LK dimers have significantly enhanced cell penetration activities
compared to those of the corresponding monomer. This feature results in near quantitative uptake of
dimeric peptides in HeLa and macrophage cells and in cell activities against TAR RNA at low
nanomolar concentrations. By using qRT-PCR and a luciferase reporter assay, we showed that the LK
dimers display dose dependent inhibition of the Tat-TAR interaction with IC50 values that are ca. 10
nM. In addition, the dimeric peptides are not cytotoxic at concentrations <2 µM, suggesting that TAR is
their dominant RNA target at low concentrations in the cells. The results of experiments using HIV-1infected cells show that the LK dimeric peptides inhibit viral replication at nanomolar concentration.
This finding strongly suggests that the LK dimers have the potential of serving as effective anti-HIV-1
drugs that function by inhibiting the Tat-TAR interaction that takes place in acute phases of the
disease.
P-14
Single Chirality Separation of Single-Walled Carbon Nanotube Using Flavin
Mononucleotide by Density Gradient Ultracentrifugation
Myungsu Jang1, Haneul Jeong1, Jinsook Sim1, and Sang-Yong Ju1*
1
Department of Chemistry, Yonsei University, Seoul 120-749, Republic of Korea
syju@yonsei.ac.kr
As-synthesized single-walled carbon nanotubes (SWNTs) are typically highly agglomerated and
heterogeneous in physical properties, thereby individual dispersion and subsequent separation of
SWNTs with single chirality and identical properties are critical steps enabling various high-end
applications such as electronic devices and biological applications. We utilize flavin mononucleotide
(FMN), derivative of riboflavin (vitamin B2), to separate a single chirality of SWNTs. When FMN wraps
SWNTs in helical pattern, hydrogen bonding between FMN moieties and π-π interaction on graphene
sheet of carbon nanotube sidewall allows tight binding in efficient manner. The separated single
chirality SWNT from FMN-suspended SWNTs were enabled by density gradient ultracentrifugation
(DGU) method, according to both sedimentation coefficients and buoyant density, related to binding
affinity between FMN surfactant and various chirality of SWNTs. Unlike other surfactant system as
sodium cholate (SC) DGU showing diameter-dependent sorting tendency, FMN DGU system displays
sorting tendency according to binding affinity to SWNTs. Since separation mechanisms of SWNTs by
DGU are not fully understood yet, we expect to reveal the DGU mechanisms by new surfactant
system. Separation of various single chirality SWNTs with high purity is undergoing.
- 57 -
P-15
Protein controlled bipolar doping of a monolayer graphene
Ji-ryang Jang a,§ , Sung Kyu Jang a,§ , Sungjoo Lee a,b,c* , Woo-Seok Choe a,d*
a
SKKU Advanced Institute of Nanotechnology (SAINT), Sungkyunkwan University, Korea
b
Center for Human Interface Nanotechnology (HINT), Korea
c
College of Information and Communication Engineering, Sungkyunkwan University, Korea
d
School of Chemical Engineering, Sungkyunkwan University, Korea
§
These authors contributed equally to this work.
*
To whom the correspondence should be addressed.
checws@skku.edu (W.-S. Choe); leesj@skku.edu (S. Lee)
In this work, p- and/or n-type doping of chemical vapor deposition-grown graphene is controlled with
the use of protein bovine serum albumin (BSA) as a dopant. BSA acts as either an electron donor or
acceptor when layered on the graphene surface in denatured form by undergoing protonation or
deprotonation reaction subject to solution pH. The direct annealing of graphene with denatured BSA
through π-stacking interaction and the subsequent pH-dependent charge modulation using a single
dopant facilitated fabrication of p- and/or n-type graphene transistor. Following AFM confirmation of
the BSA/graphene interface assembly, the carrier transport properties of BSA-doped graphene
transistors were assessed by I-V measurement and Raman spectra to show effective charge
modulation of the BSA-doped graphene at various pH conditions. In conclusion, the protein-mediated
bipolar doping of graphene demonstrated in our study is simple, scalable and straightforward, thereby
providing a useful alternative for fabricating graphene transistors of novel properties.
P-16
Graphene Nanoribbons Formed by a Sonochemical Method using Flavin
Mononucleotide Assembly as a Template
Sang-Yong Ju* and Woojin Yoon
Department of Chemistry, Yonsei University, Seoul 120-749, Republic of Korea
syju@yonsei.ac.kr
Graphene with nanotexture can be a
valuable way to control its electronic and
physicochemical property. We show that
flavin mononucleotide (FMN)-assisted
dispersion produce few-layered graphenes,
including nanoribbons and nanocuts. The
dispersion displayed a high optical density
up to 0.24 mg/mL of graphene. Raman
spectroscopy revealed that the graphene
possesses large disorder-related bands (D
and D‟ bands), along with the G and D bands, and exhibits an intensity ratio of D over D‟ bands near 4,
suggesting lined defect. Such behavior was confirmed by high-resolution transmission electron
microscopy (HRTEM), where graphene flakes contained dense and straight graphene cuts and
nanoribbons less than a few tens of nanometers in width. Such ribbon formation originated from
sonochemical graphene unzipping using a one-dimensional FMN ribbon as a template. The defect
density measured by HRTEM and Raman studies suggested that a single line defect per 7-15 nm.
Graphene films with 75% transmittance displayed a low sheet resistance (28 k/sq) better than that of
reduced graphene oxide (RGO). This study provides a valuable tool to produce graphene nanoribbons,
and solution-processable graphene dispersions to future applications.
- 58 -
P-17
Development of pancreatic β-cell probes binding to insulin for in vivo imaging
Nam Young Kang1, Jungyeol Lee2, Sung-Jin Park 1, Wut Hmone Phue2, Bikram Keshari
Agrawalla2, Young-Tae Chang1,2
1
Laboratory Bioimaging Probe Development, Singapore Bioimaging Consortium, Singapore
2
Department of Chemistry, National University of Singapore, Singapore
Kang_nam_young@sbic.a-star.edu.sg
The insulin-producing beta cells within the islets of the pancreas are essential for maintaining glucose
homeostasis. Because of this, the pancreatic beta cells are a major target for the treatment of diabetes
as well as imaging of viable pancreatic islet beta cells is an important component in research on
diabetes both in clinical and experimental medicine. The way of potential cure patients with type 1
diabetes is to do pancreatic islet transplantation that is has emerged as an effective therapy. Since
diabetic patients only need the pancreatic islets, which contain the insulin-secreting beta cells, the
idea of separating the islets and transplanting only them is very attractive. One of major obstacle to
islet transplantation is how to monitor the pancreatic healthy islets and measure insulin secretion after
transplantation using reasonable non invasive methods. We have recently reported the small molecule
probe for selective staining of beta cells in pancreatic islet for live animals. However, the probe is not
insulin target. Our goal is to identify even more specific beta cell probes binding to insulin. The key
technology to achieve this is our Diversity Oriented Fluorescence Library Approach (DOFLA), which
provide thousands of highly versatile fluorescent small molecules. Based on our powerful high
throughput screening system, we discover novel fluorescent beta cell probes for detecting pancreatic
islet and measuring insulin response. Eventually, we apply them for molecular imaging of monitoring
pancreatic beta cell of islets after transplantation. In this point of view, our small molecule beta cell
probes having inherent fluorescence can be used as direct optical imaging probes as isolation tool to
purify the islets of sufficient mass. This capability will provide an enormous advance in our prediagnosis and care of diabetes patients while generating novel approaches to develop better diagnosis
against diabetes.
P-18
A New Genetically Encodable Flavinylation Tag for Screening Proteasome Inhibitors In
Living Mammalian Cells
Myeong-Gyun Kang and Hyun Woo Rhee*
Department of Chemistry, Ulsan National Institute of Science and Technology (UNIST), Korea
rukia123@unist.ac.kr
Recently, it was found that ApbE, a bacterial flavin transferase, catalyzes flavin adenosine
dinucleotide (FAD) and transfers Flavin mononucleotide (FMN) to a threonine residue (Thr-229) in
bacterial flavoprotein, nqrC (Bogachev, J. Biol. Chem., 2013). Because FMN has considerable strong
green fluorescence (λem= 525nm, Q=0.26), it allows FMN-conjugated protein could be detected by
fluorescence detection methods. Herein, we firstly try to see flavin transferase reaction between ApbEnqrC in mammalian cell by recombinant protein expression. In result, it showed very orthogonal
flavinylation reaction. Also, we found that the nqrC targeted to endoplasmic reticulum lumen is
degraded by cytosolic proteasome through unfolded protein response (URP) pathway. We used MG132 which is a proteasome inhibitor for reducing the degradation of nqrC by the proteasome and then
flavinylated nqrC level in cytosol increases as concentration of MG-132 increases. This result allows
potential fluorescence-based high-throughput screening of the drug candidates related to proteasome
inhibition, which has potential for anti-cancer treatment.
- 59 -
P-19
Sending innate immune signals across the membrane: A multiscale simulation
approach to Toll-like receptor assembly
Vasileios E Kargas1,3, Daniel A Holdbrook1, Isaac Godfroy4, Hang H Yin4, Robert C Ford3, Peter J Bond1,2
1 Bioinformatics Institute (A*STAR), 30 Biopolis Str, #07-01 Matrix, Singapore 138671.
2 Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, 117543 Singapore.
3 Faculty of Life Sciences, The University of Manchester, Dover Str, M139PT, United Kingdom.
4 Department of Chemistry and Biochemistry and the BioFrontiers Institute, University of Colorado Boulder,
Boulder, CO 80309, USA
Vasileiosk@bii.a-star.edu.sg
Toll-like receptors (TLRs) are single transmembrane-spanning proteins that sense pathogenic
molecular patterns within the innate immune system. Upon activation, TLRs form homodimers or
heterodimers and initiate immune response pathways. The ability of TLRs to dimerize is therefore
critical to their function in responding to invading pathogens. Crystallographic structures have been
solved for the ectodomains of various TLR homodimers, but high-resolution structural information is
not available for the full-length proteins, or for the transmembrane (TM) regions. We performed abinitio modelling of the TM regions of all ten human TLRs, based on secondary structural predictions
and spectroscopic data. Subsequently, coarse grained (CG) molecular dynamics (MD) simulations
were performed to follow assembly and homo/hetero-dimerization within a phospholipid membrane
environment. Our results have been used to evaluate the stability of TLR dimers, and to identify key
sequence motifs that stabilize TM interactions, helping to rationalize in vitro data. Using acceptor
photobleaching FRET on live cells, it has been demonstrated that TM domains including TLR2-TLR1
and TLR2-TLR6 interact within the plasma membrane. Additionally, multiscale models of entire TLR
receptors have been built to determine the link between ligand recognition, assembly and crossmembrane/downstream signaling. In particular, we have focused on TLR4, which recognizes
lipopolysaccharide (LPS) from the outer membranes of Gram-negative bacteria, for which a variety of
structural/biophysical and MD data are available. Modelling of mutant constructs containing variable
linkers revealed the structural basis for experimentally demonstrated tight coupling between extra- and
intra-cellular domains and the TM region, based on receptor stability and dimerization efficiency.
These data improve our understanding of the assembly and signalling mechanisms in TLRs, and may
facilitate design of ligands with specific immunomodulatory properties, paving the way for new
therapeutic treatments of inflammatory diseases.
P-20
Synthesis and Binding Analysis of Thioflavin T Analogs Targeting G-Quadruplex
1
1
1,2
1
1
Yuka Kataoka , Hiroto Fujita ,Yuuya Kasahara , Toshitada Yoshihara , Seiji Tobita , Masayasu Kuwahara
1
1
Graduate School of Science and Technology, Gunma University, Japan
2
National Institute of Biomedical Innovation (NIBIO), Japan
mkuwa@gunma-u.ac.jp
Small molecular ligands that specifically bind to target biomolecules have been extensively studied for
their applications as molecular probes and drugs. For example, an amyloid-binding fluorescent dye,
Thioflavin T (ThT), has been used as a small molecule ligand. ThT is likely a unique ligand, particularly
1
because it maintains protein homeostasis and extends the lifespan of Caenorhabditis elegans.
Furthermore, ThT can also stabilize the antiparallel G-quadruplex form of 22AG human telomeric DNA
2
and enhance its fluorescence emissions. DNA secondary structures of a G-quadruplex are formed
with certain G-quartets that are four Hoogsteen-paired coplanar guanines. Some G-quadruplex
topology types have been reported thus far: parallel, antiparallel, hybrid1, and hybrid2 (mixed). These
sequences are frequently observed near transcription start sites in a gene promoter and its 5′-UTR
elements as well as in telomeric regions. Therefore, it has been proposed that these G-quadruplexes
may have important roles in cellular processes, including transcription. These reports also suggest
that G-quadruplex-binding ThT analogs may be able to control gene expression.
Therefore, in this study, we synthesized novel ThT analogs and investigated the binding properties of
G-quadruplex-binding ThT and ThT analogs under physiological salt conditions. We observed that
these ThT analogs could distinguish between different G-quadruplex topology types based on their
fluorescence emissions.
1. Alavez, S.; Vantipalli, M. C.; Zucker, D. J.; Klang, I. M.; Lithgow, G. J. Nature 2011, 472, 226–229.
2. Mohanty, J.; Barooah, N.; Dhamodharan, V.; Harikrishna, S.; Pradeepkumar, P. I.; Bhasikuttan, A.
C. J. Am. Chem. Soc. 2013, 135, 367–376.
- 60 -
P-21
Identification of prion-like tau species
Dohee Kim, Sung-su Lim, Mamun Harque, Yun Kyung Kim*
1
Korea Institute of Science and Technology (KIST), Brain Science Institute, Center for neuromedicine, Seoul 136-791, South Korea
2
Biological Chemistry, University of Science and Technology (UST), Daejeon 305–333, South Korea
yunkyungkim@kist.re.kr
Abnormal tau aggregation is a pathological hallmark in multiple neurodegenerative diseases
collectively called tauopathies. Mounting evidences suggest that tau aggregates are toxic to neurons,
but also propagate in neurons acting as a seed for native tau aggregation. Accordingly, prion-like tau
transmission become an important pathogenic mechanism driving the disease progression. Although
its pathological importance, prion-like tau species are not fully characterized yet. To identify the
pathogenic tau species, we prepared diverse tau aggregates and compared their prion-like activity in
tau-BiFC cells, a cell-based senor for monitoring tau-tau interaction in a range of dimers to large
aggregates. Among tested, soluble forms of P301L mutant tau actively increased intracellular tau
assembly and phosphorylation. Further biochemical analysis of the pathogenic tau species identified
that P301L mutant tau prefers to form structurally stable dimes and trimmers, which are conjugated by
intermolecular disulfide cross-links. Full-length human tau (441 a.a.) contains only two cysteine
residues that can form both intra- and inter-molecular disulfide bonds. Our results suggest that the
disulfied cross-inkage serves structural supports for tau oligomers that are critical for
neurodegeneration and transmission of the disease.
P-22
Peptide-mediated synthesis of palladium raspberry nanoparticles and their effect on
catalytic activities in Sonogashira cross-coupling reaction
Young-O Kim1, Hyeong-Seok Jang1, Yo-Han Kim1, Jae Myoung You1, Yoon-Sik Lee1
1
School of Chemical and Biological Engineering, Seoul National University, Korea
zerofive@snu.ac.kr
Nanomaterials have received a lot of attention because of various properties depending on their size,
morphology, and atomic arrangement. To control these properties, a number of methods have been
tried using various stabilizer as well as kinetic conditions. In biological organisms including sponges,
mollusks, and diatoms, inorganic nanostructures are manipulated through bio-mineralization
processes under physiological conditions. For this process, three dimensional structure of polypeptide
subunit is important to grow inorganic or metal nanoparticles (NPs). Recently, to mimic this system,
self-assembled peptides have been focused to synthesize metal NPs. Here, tyrosine-rich peptides
(TRPs) are chosen as a template to mimic the bio-mineralization. We have found that peptide
+2
assembly can be induced through interactions between TRPs and palladium (Pd ) ions. During this
process, tyrosine residues might afford specific folding geometry and regulate its surface energy.
Therefore, peptides can control the uniform growth of raspberry-shaped Pd NPs (4-5 nm diameter) by
peptide ligands. In addition, these raspberry-shaped Pd NPs showed good catalytic activities in
copper-free Sonogashira cross-coupling reaction in water.
- 61 -
P-23
Efficient aerobic oxidation of alcohols to aldehydes over graphene oxide supported
ruthenium oxide catalyst under aqueous condition
Yo-Han Kim1, Jung Won Kim2, and Yoon-Sik Lee1
1
2
School of Chemical and Biological engineering, Seoul National University, Seoul, 151-742, Republic of Korea
Department of Chemical Engineering, Kangwon National University, Samcheok, 245-711, Republic of Korea
yslee@snu.ac.kr
The aerobic oxidation of alcohols to the corresponding carbonyl compounds is important in organic
chemistry, because it brings several advantages such as free of environmentally undesirable solvent
and heavy-metal waste, and wide application of their oxidized products as various chemical
intermediates. Thus, molecular oxygen has been received much attention as an alternative of
stoichiometric oxidant. However, it is hard to activate molecular oxygen for aerobic alcohol oxidation
because of high activation barrier and low solubility of molecular oxygen in water. Thus, developing
water compatible catalyst is crucial issue for aqueous aerobic alcohol oxidation. Here, we developed
the GO supported ruthenium oxide catalyst (RuO 2/GO), which showed excellent catalytic performance
in water for the oxidation of various alcohol substrates with molecular oxygen with TON value of up to
10,000. The RuO2/GO catalyst could be reused without significant loss of its catalytic activity.
P-24
Crystallographic and biochemical studies of ilvC, a ketol-acid reductoisomerase, from
Streptococcus pneumoniae D39
GyuHee Kim1, GyuLee Kim2, Sumin Lee3, Jaesook Yoon3, Dong-Kwon Rhee2,
and Sangho Lee1
1
Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746, Korea
2
School of Pharmacy, Sungkyunkwan University, Suwon 440-746, Korea
3
Department of Chemistry, Sungkyunkwan University, Suwon 440-746, Korea
sangholee@skku.edu
Streptococcus pneumoniae is a gram-positive bacterium causing respiratory diseases in human.
Despite extensive studies in characterizing virulence factors in S. pneumoniae, the emergence of
multi-drug resistant strains and new serotypes demands search for novel virulence factors.
Biosynthesis of branched-chain amino acids (BCAA) is reportedly required for survival and virulence of
a bacterial pathogen. To investigate whether the BCAA biosynthetic pathway is involved in
pneumococcal growth and virulence, we studied ilvC from S. pneumoniae D39 (SpIlvC), a ketol-acid
reductoisomerase. SpIlvC catalyzes the second step in biosynthesis of branched-amino acids (BCAA)
such as isoleucine, leucine, and valine. We determined the crystal structure of SpIlvC at 1.69 Å
resolution. The structure of SpIlvC contains an asymmetric dimer in which one subunit is in apo form
2+
and the other in NADP(H) and Mg -bound form. Structural analysis combined with activity assay
using site-directed mutants suggested that Asp-83 in the NADP(H) binding site and Glu-195 in the Mgcoordination site are the most critical in the activity of SpIlvC. To investigate local conformational
changes in the active site mutants, we determined the crystal structures of D83G and E195S at 2.29
and 2.02 Å resolution, respectively. Superposition of wild-type with D83G and E195S structures
2+
revealed that the local conformational change is observed in Mg -coordination and NADP(H) binding
region with r.m.s.d. being 3.20 and 3.17 Å for D83G and E195S, respectively. When the gene for
SpIlvC was disrupted, pneumococcal growth was retarded and virulence attenuated in vivo. Taken
together, our results demonstrate that SpIlvC is critical in pneumococcal growth and possibly involved
in the virulence.
- 62 -
P-25
Exploitation of chemistry and biology of siderophores to combat against drugresistant bacteria
Hak Joong Kim, Hwisoo Ree, Jimin Kim, Hyeon Seok Kim, Woon Young Song, Seong Ji
Choi, Mijin Sun, Jin-su An, Jae Eun Lee
Department of Chemistry, Korea University, Seoul 136-701, Republic of Korea
hakkim@korea.ac.kr
The occurrence and world-wide spread of drug-resistant bacteria, often referred as “superbacteria”,
have emerged as significant threats in recent years. However, despite their significant adverse
impacts on human welfare, only a handful of new antibiotic drugs have been introduced to clinics over
the past decades, but, unfortunately, acquisition of the resistance to those agents by pathogens has
been already reported. Thus, there is an urgent need to discover effective antibiotic agents of which
mode of action is distinctive from those of traditional drugs to overcome the resistance problem.
Siderophores are small molecule chelators produced and utilized by most pathogens for assimilation
of the iron from the environments. Due to the extremely low bioavailability of the iron in the infected
hosts, siderophore-related mechanisms serve as important virulent factors, and thus inhibitions thereof
have been considered as viable targets for novel antibiotic discovery. In addition, application of
siderophores as drug-delivery vectors has also attracted great attentions, because the cellular entry of
siderophores utilizes designated uptake machineries independent of porin-based passive diffusion.
Therefore, such “Trojan horse” strategy could be particularly effective against pathogens of resistance
was associated with reduced membrane permeability. In this regard, this presentation describes
various efforts of our laboratory to fully exploit this novel approach for antibiotic discovery, particularly
focusing on the establishment of chemical syntheses of a number of siderophores as well as
development of novel fluorescence probes to quantitatively assess cytoplasmic uptake of siderophoredrug conjugates.
P-26
Harnessing Peptides for Electrochemical Detection of Bisphenol A
Sung-Eun Kim1, Jiao Yang1, Misuk Cho1, Ik-Keun Yoo2, Youngkwan Lee1,*,
Woo-Seok Choe1,*
1
School of Chemical Engineering, Sungkyunkwan University, Republic of Korea
School of Chemical Engineering and Bioengineering, University of Ulsan, Republic of Korea
walkyrie86@skku.edu
2
Bisphenol A (BPA), an organic compound with two phenol functional groups, is widely used for the
production of plastics, adhesives, flame retardants and it is released into the environment primarily via
wastewater during the manufacturing process or by leaching from commercial plastic products. BPA
has been implicated as an endocrine disruptor that is capable of mimicking natural hormones, thereby
leading to heart diseases, diabetes and cancer. Thus, fast and accurate detection of BPA in varying
matrices has become important in recent years. In this study, a cysteine-flanked heptapeptide
sequence Cys-Lys-Ser-Leu-Glu-Asn-Ser-Tyr-Cys (CKSLENSYC), which is capable of recognizing
BPA with high specificity, was isolated using a phage display technique. A novel electrochemical
biosensor harnessing this affinity peptide as a BPA detection probe was constructed and its
performance was assessed. The affinity peptide was immobilized onto a gold electrode via Au-S
bonding and the formation of self-assembled peptide monolayer on the electrode was confirmed by
attenuated total reflection infrared spectroscopy (ATR-IR), cyclic voltammetry (CV) and
electrochemical impedance spectroscopy (EIS). In order to maximize the performance of the
constructed sensor on BPA detection, the effects of some critical factors (i.e., detection time and pH
condition) on probe-target recognition were also assessed. Following the exploration of the optimum
sensing condition, differential pulse voltammetry (DPV) was used to determine the varying
concentrations of BPA in the solution. The developed sensor conveyed excellent performance in view
of sensing speed, sensitivity and selectivity by detecting BPA in less than 5 min with a broad dynamic
detection range of 1-5000 nM of BPA, despite the presence of several interfering species, such as
phenolic compounds and inorganic ions.
- 63 -
P-27
Engineering of Mono-Disperse Oligoethylene Glycols for Protein Manipulation
K. Kinbara*1, T. Muraoka1, K. Adachi1, M. Ui1, S. Kawasaki1, N. Sadhukhan1,
H. Obara1, M. Laguerre2, H. Tochio3 and M. Shirakawa3
1
Institute of Multidisciplinary Research for Advanced Materials, Tohoku University
2
Institut européen de chimie et biologie
3
Department of Molecular Engineering, Graduate School of Engineering, Kyoto University
kinabra@tagen.tohoku.ac.jp
Polyethylene glycol (PEG) is one of the most popular water-soluble organic compounds, which has
been widely used for biological and medicinal applications. Nonionic, water-soluble and nontoxic
characters of PEG are particularly useful for pharmaceutical applications. Covalent attachment of
PEG chains to a target molecule such as a drug or a therapeutic protein, is know as “PEGylation”, that
inhibits protein aggregation and prolongation of drugs‟ circulatory time. However, it is also known that
linear PEG chains tend to cover the surface of proteins to cause serious deactivation of their functions.
Here we propose novel molecules composed of ethylene oxide skeleton, as “structured” PEG
molecules, that adopt rigid 2D or 3D structures and hold the merits of PEG. In order to develop
structured PEGs having 2D or 3D molecular shapes, we have investigated synthetic methods to
connect several tetraethylene glycol (TEGs) units with pentaerythritol. As the first example of the
structured PEG, we have developed macrocyclic triangle PEG that consists of three TEG and
pentaerythritol units, respectively.
Interestingly, this triangle molecule exhibits hydrophilicity/hydrophobicity switching at lower
temperature than the linear PEG with a comparable molecular weight, and effectively suppresses
thermal aggregation of lysozyme.
References
1. T. Muraoka, K. Adachi, M. Ui, S. Kawasaki, N. Sadhukhan, H. Obara, H. Tochio, M. Shirakawa and
K. Kinbara, Angew. Chem. Int. Ed., 2013, 52, 2430.
2. T. Muraoka, N. Sadhukhan, M. Ui, S. Kawasaki, E. Hazemi, K. Adachi and K. Kinbara, Biochem.
Eng. J., 2014, 86C, 41.
3. T. Muraoka, K. Adachi, R. Chowdhury and K. Kinbara, PLoS ONE, 2014, 9, e91912
P-28
Stylissatin A, A Cyclic Peptide That Inhibits Nitric Oxide Production
from Marine Sponge
Masaki Kita, Baro Gise, Atsushi Kawamura, Taiki Sunaba, Tito Akindele, Hideo Kigoshi
Graduate School of Pure and Applied Sciences, University of Tsukuba, Japan
mkita@chem.tsukuba.ac.jp
Inflammation is a cellular and vascular response that occurs immediately after tissue damage. Upon
exposure to various incoming signals from the cell surface, a series of proteins are activated in
phagocytes, and the free radical nitric oxide (NO) is produced by nitric oxide synthases (NOS). The
purpose of this study is to identify pharmacological agents that can regulate NO production in
phagocytes, which may contribute to the development of new anti-inflammatory agents. In our
continuing search for bioactive substances from marine invertebrates, we have isolated a new cyclic
heptapeptide, stylissatin A, from the Papua New Guinean marine sponge Stylissa massa. Through the
use of 1D and 2D NMR spectroscopic analysis, Marfey‟s method, and MS/MS analysis, its structure
1
2
3
4
5
6
7
was determined to be cyclo-[Tyr –Ile –Phe –Pro –Ile –Pro –Phe ]. Stylissatin A inhibited NO
production in LPS-stimulated murine macrophage RAW264.7 cells with an IC 50 value of 87 µM. By
using solution and solid-phase peptide synthesis, we have synthesized stylissatin A and established
its absolute configuration and biological activity. We will present the chemical and biological studies of
this unique heptapeptide and its synthetic derivatives.
- 64 -
P-29
Modified DNA Aptamers that Bind to Vascular Endothelial Growth Factors
Naoto Honda,1 Kenta Hagiwara,1 Yuuya Kasahara,1,2 Masayasu Kuwahara1,*
1
Graduate School of Science and Technology, Gunma University, Japan
2
National Institute of Biomedical Innovation (NIBIO), Japan
mkuwa@gunma-u.ac.jp
Vascular endothelial growth factors (VEGFs) are a group of glycoproteins that are involved in
vasculogenesis, neovascularization, angiogenesis, and tumor metastasis. Thus, VEGF-binders such
as bevacizumab and pegaptanib have been developed as therapeutic agents. As with pegaptanib,
nucleic acid aptamers are promising candidates for medicines; therefore, various nucleic acid
aptamers for a broad range of protein targets are now being developed. It was recently found that
high-affinity aptamers for so-called difficult proteins, for which high-affinity aptamers could not
generally be obtained from natural DNA/RNA libraries using conventional systematic evolution of
ligands by exponential enrichment methods, could be acquired when using certain types of basemodified DNA libraries.
Therefore, in this study, we prepared modified DNA aptamers that could specifically bind to VEGF-165
using a capillary electrophoresis–systematic evolution of ligands by exponential enrichment (CE–
SELEX) method with two different base-modified DNA libraries. Aptamer selection from a natural DNA
library was also performed using similar procedures and conditions.
1. (a) J.D. Vaught, C. Bock, J. Carter, T. Fitzwater, M. Otis, D. Schneider, J. Rolando, S. Waugh, S.K.
Wilcox, B.E. Eaton, J. Am. Chem. Soc., 2010, 132, 4141–4151. (b) L. Gold, D. Ayers, J. Bertino, C.
Bock et al., PLoS One, 2010, 5, e15004. (c) D.R. Davies, A.D. Gelinas, C. Zhang, J.C. Rohloff,
J.D. Carter, D. O'Connell, S.M. Waugh, S.K. Wolk, W.S. Mayfield, A.B. Burgin, T.E. Edwards, L.J.
Stewart, L. Gold, N. Janjic, T.C. Jarvis, Proc. Natl. Acad. Sci. USA, 2012, 109, 19971–19976. (d)
Y. Imaizumi, Y. Kasahara, H. Fujita, S. Kitadume, H. Ozaki, T. Endoh, M. Kuwahara, N. Sugimoto,
J. Am. Chem. Soc., 2013, 135, 9412–9419.
2. (a) Y. Kasahara, Y. Irisawa, H. Fujita, A. Yahara, H. Ozaki, S. Obika, M. Kuwahara, Anal. Chem.,
2013, 85, 4961–4967. (b) Y. Kasahara, Y. Irisawa, H. Ozaki, S. Obika, M. Kuwahara, Bioorg. Med.
Chem. Lett., 2013, 23, 1288–1292. (c) M. Kuwahara, S. Obika, Artif. DNA PNA XNA, 2013, 4, 39–
48.
P-30
Exploring the Interaction of Quantum Dots Hybrid with Cells
San Kyeong, Cheolhwan Jeong, Bong-Hyun Jun and Yoon-Sik Lee
School of Chemical & Biological Engineering, Seoul National University
feanor05@snu.ac.kr
For decades, the nanoparticle (NP) based materials have been actively studied in various fields of
biomedical and clinical research. However, the interactions between the physico-chemical surface
properties of the NPs and their adsorption/uptake tendencies by living cells have been poorly
understood. Herein, in order to expand the knowledge on the correlation between the surface
chemistry of NPs and their cell penetration property, we prepared a set of QD-embedded silica NPs
(SiO2@QDs@SiO2 NPs), which have different kinds of surface charges and measured their adsorption
and uptake tendencies by various kinds of cells.
- 65 -
P-31
Fluorescence Probe Development for Lung Cancer Stem Cells
Yong-An Lee1, Srikanta Sahu2, Animesh Samanta2, Hwa-Young Kwon3, Nam-Young Kang1,
Young-Tae Chang1,2
1
Laboratory Bioimaging Probe Development, Singapore Bioimaging Consortium, Singapore
2
Department of Chemistry, National University of Singapore, Singapore
3
Singapore Centre on Environmental Life Sciences Engineering, Singapore
Lee_yong_an@sbic.a-star.edu.sg
A growing body of evidence suggests that cancers are organized with a subpopulations of cancer
stem cells (CSCs) that generate the tumor by propagating growth and metastasis of tumor cells. CSCs
are thought to be responsible for cancer relapse after difinitive therapy by a preferential resistance to
cancer drugs. Therefore, understanding of CSCs may lead to crucial advances in more effective
therapies. Lung cancer is the leading cause of cancer death worldwide, with less than 15% of patients
surviving beyond 5 years. The possibility to characterize biological properties of lung CSCs may
provide powerful tool to improve the clinical outcome of lung cancer patients. As the result of efforts to
understand lung CSCs, the previous study shows that CD166 can be used for isolation of lung CSCs
+
and CD166 fraction has ability to form tumor spheres, a widely used in vitro technique for assessing
self renewal capacity of CSCs. Thus, lung tumor sphere (TS) cell line, which is enriched for lung CSCs,
can be established by using the CD166 antibody.
The aim of the present study is to develop the fluorescent probes for isolation and in-vivo detection of
lung CSCs. The key technology to achieve this is our Diversity Oriented Fluorescence Library
Approach (DOFLA), which provide thousands of fluorescent small molecules. Screening of the
fluorescent small molecules with lung TS cells identified two types of fluorescent probes of visible
spectrum (VIS) and near infra-red (NIR) spectrum, respectively, that selectively labels lung TS cells.
We, eventually, confirmed that both compounds are can be applied for isolation of lung CSCs from
tumor.
In conclusion, this capability will provide an enormous advance in cancer treatment and care of
patients through novel approaches to discover molecular markers and in vivo imaging probe
development for lung CSCs using the VIS and NIR probes.
P-32
APEX-Generated „Barcode‟ Tells Spatial Information of Target Protein in Living
Mammalian Cells
Song-Yi Lee and Hyun Woo Rhee*
Department of Chemistry, Ulsan National Institute of Science and Technology (UNIST), Korea
song@unist.ac.kr
In human cell, there are 18,000 different kinds of protein. Each protein is localized into specific
subcellular compartments. Localization information of certain protein is important because it tells many
things about its functions. However, optical imaging method cannot achieve to get sub-organelle
localization information due to its low resolution power (> 220 nm). For identification of sub-organelle
localization information, there are developed some methods such as electron microscope (EM)
imaging or high-resolution mass spectrometry. However, those methods require expensive equipment,
trained researchers, and laborious preparation steps. Herein, we propose a simple pattern-based
method to identify protein subcellular localization by using engineered ascorbate peroxidase (APEX)
with western blotting (WB) method which is generally performed in most of cellular biology laboratories.
For endogenous proteome inventory of each organelle is very different from each other, we found
biotinylated proteins by each organelle targeted APEX were also very different. Interestingly, APEXs‟
in same sub-organelle with different targeting sequences showed very similar western blotting pattern
by streptavidin-HRP. We believe these distinct APEX-generated WB patterns of various compartments
works as authentic „barcode‟ to get meaningful spatial information of protein of interest in suborganelle resolution.
- 66 -
P-33
Proteome reactivity profiling for the discrimination of pathogenic bacteria
Jun-Seok Lee*
Molecular Recognition Research Center, Korea Institute of Science and Technology (KIST), South Korea
junseoklee@kist.re.kr
Activity-based protein profiling (ABPP) has been successfully applied to monitor the functional state of
enzymes in complex proteomes using reactive chemical probes. The basis of ABPP is the use of
chemical probes in which the electrophiles are connected to reporter tags. Chemical probes form
covalent bonds at the active site of enzymes via a catalytic reaction, and the labeled enzymes can be
further visualized or identified in a high-throughput manner. Depending on the reactive functional
groups, many kinds of enzymes can be targeted, including serine proteases, cysteine hydrolases, and
kinases. Series of electrophiles have been extensively investigated as chemical probes to unveil target
proteins and their preference for labeling amino acid residues. Based on the previous reports, smallmolecule electrophiles often lack selectivity toward a single class of enzyme superfamily, and they can
target distinct classes of enzymes instead. Moreover, a few hyper-reactive residues can also make a
covalent bond, though such residues are not present at the enzymatic active site. We envisioned that
all kinds of reactivity profile could be a valuable property to generate fingerprints of the individual
pathological status of proteomes.
P-34
A Facile Method for Sequence Determination of Cyclic Peptides/Peptoids via CNBrMediated One-Pot Ring-Opening/Cleavage Reaction
Kang Ju Lee, Hyun-Suk Lim*
Department of Chemistry, Pohang University of Science and Technology, Pohang 790-784, Korea.
kjulee0130@gmail.com
Macrocyclic peptides and peptidomimetics (e.g., cyclic peptoids) are of enormous interest as a
promising class of protein binding ligands. Compared to their linear counterparts, cyclic peptides and
peptidomimetics have increased conformational rigidity and relatively pre-organized structures, thus
could bind more tightly to the target proteins without a major entropy loss. Moreover, they often have
improved cell permeability and proteolytic stability. Owing to their relatively large size and
conformational constraint, macrocyclic peptides and peptidomimetics might be well-suited molecules to
target many challenging protein targets such as protein-protein interactions (PPIs), which cannot be
easily modulated by traditional drug-like small molecules.
Despite many favorable features, the utility of macrocyclic peptides and peptidomimetics is limited by
challenges in their sequence determination when using them in high-throughput screening (i.e. onbead screening). Cyclic peptides and peptidomimetics from one-bead one-compound (OBOC)
combinatorial libraries are not generally sequenced by Edman sequencing and tandem mass
spectrometry. Here, we designed a cyclic system for sequencing cyclic peptides and peptoids, which
has a thioether moiety embedded in the backbone of them. The thioether linkage is cleaved via a fivemembered ring intermediate by treating with CNBr through the similar mechanism by which CNBr
hydrolyzes methione-containing peptide sequences. Subsequent hydrolysis of the intermediate
imminium salt results in cleavage of the linearized peptides/peptoids from the bead. The sequence of
the resultant linear peptides/peptoids can be analyzed by tandem mass spectrometry.
- 67 -
P-35
Layer-by-Layer-Based Silica Encapsulation of Individual Yeast
Hojae Lee1, Daewha Hong1, Taegyun Park1, Hee Chul Moon1, Insung S. Choi1
Center for Cell-Encapsulation Research and Molecular-Level Interface Research Center, Department
of Chemistry, KAIST, Korea
ischoi@kaist.ac.kr
In the area of cell-surface engineering with nanomaterials, the metabolic and functional activities of the
encapsulated cells are manipulated and controlled by various parameters of the artificial shells that
encase the cells, such as stiffness and elasticity, thickness, and porosity. The mechanical durability
and physicochemical stability of inorganic shells prove superior to layer-by-layer-based organic shells
with regard to cytoprotection, but it has been difficult to vary the parameters of inorganic shells
including their thickness. In this work, we combine the layer-by-layer technique with a process of
bioinspired silicification to deposit functional nanoparticles inside the silica shells, and to control the
thickness of the silica shells.
P-36
Dual-labeled multivalent glycoconjugates for detection of cell-surface lectins
Hui Li1, Xizhe Tian1,2, Kyung-Hwa Baek1 and Injae Shin1
1
National Creative Research Center for Biofunctional Molecules, Department of Chemistry, Yonsei
University, Seoul 120-749, Korea
2
Department of Chemistry, College of Sciences, Yanbian University, Yanji 133000, China
injae@yonsei.ac.kr
To date, many efforts have been made to detect lectins in cells by using single imaging techniques.
Lectin binding properties of the glycoclusters were initially examined by using microarrays immobilized
by various lectins. These glycoclusters were then employed to detect the cell-surface carbohydratebinding proteins in bacteria. Moreover, the uptake of glycoclusters by mammalian cells through
receptor mediated endocytosis was evaluated. However, only a few dual-labeled glycan-based probes,
which integrate advantageous features of two imaging methods to enhance the visualization of
biological processes associated with lectins in cells, have been reported. Herein we describe the
synthesis of dual fluorescence and magnetic resonance imaging agent conjugated neoglycopeptides
and their application in the simultaneous imaging of lectins in mammalian cells. The dual-labeled
neoglycopeptides bind to lectins on cell surfaces and subsequently enter the cells via lectin-mediated
endocytosis. The results of these efforts show that the novel dual-labeled neoglycopeptides are
effective fluorescence and MR imaging agents for monitoring biological processes associated with
lectins.
- 68 -
P-37
Structural Basis Underlying the Binding Specificity of Human Galectins-1, -3 and -7 for
Galβ1-3/4GlcNAc
Tung-Ju Hsieh, Hsien-Ya Lin, Chuen-Jiuan Huang and Chun-Hung Lin
Institute of Biological Chemistry, Academia Sinica
Academia Road Section 2, Taipei, 11529, Taiwan
pipis_lsy@hotmail.com
Galectins are β-galactoside-binding proteins that are characteristic of having one or two conserved
carbohydrate recognition domains (CRDs). Members of this family have been shown to involve in
diverse biological functions, such as cell adhesion, cell growth regulation, and apoptosis via their
interactions with β-galactoside-containing structures on cell surface. More importantly, human
galectins act as regulatory factors in many types of cancers by either inhibiting or promoting tumor
growth. Therefore, to identify selective ligands for human galectins provides not only a useful tool for
dissecting how each galectin member correlates with cancer progression, but also a possible solution
for the development of clinical therapeutics.
All galectins are able to recognize two isomers of N-acetyllactosamine, namely type 1 and 2 LacNAc
(Galβ1-3/4GlcNAc, abbreviated as LN1 and LN2, respectively) disaccharides that appear in a myriad
of glycoconjugates. Galectins share major structural homology in their CRDs, which underlies the
common binding feature with LN1- and LN2-containing glycans. Despite several crystal structures of
different galectin/LN2 complexes available, to our surprise, there is no report regarding to the structure
of galectin/LN1 complex. We herein report the crystal structures including full-length hGal1, 7 and
CRD domain of hGal3 in complex with LN1. In comparison with the analogous LN2-complexed
structures, the results pinpointed a unique Asp/Glu-Arg motif that determines the LN1- or LN2-binding
preferences of hGal1, 3 and 7, which was supported by the altered binding specificity of several sitedirected mutants. The resulting outcome sheds new insight into the different LN recognition modes of
hGal1, 3 and 7, as well as provides useful information in the development of galectin inhibitors with
improved potency and selectivity.
- 69 -
P-38
DNA structure and stability under molecular crowding conditions
with cationic polymers
Daisuke Miyoshi1, Yu-mi Ueda1, Naohiko Shimada2, Shu-ichi Nakano1, Naoki Sugimoto1,3,
Atushi Maruyama2
1
Faculty of Frontiers of Innovative Research in Science and Technology (FIRST), Konan University, Japan
2
Department of Biomolecular Engineering, Tokyo Institute of Technology, Japan
3
Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Japan
miyoshi@center.konan-u.ac.jp
One of the most distinguishing features of the intracellular environment is the condition of being
crowded with biomolecules, referred to as “molecular crowding”. There is increasing evidence that
molecular crowding has significant effects on properties of nucleic acids, and the effects are not
always the same as in the case of proteins (1,2).
Moreover, eukaryotic nuclei, where DNA exists, are highly heterogeneous and crowded with a variety
of subnuclear structures and proteins, including histones. Nuclear DNA is highly condensed and
wrapped around histone proteins, forming the nucleosomes. Such molecular environment is totally
different from a test tube condition and even from a molecular crowding condition with neutral
polymers, which are generally used for molecular crowding studies. Since the DNA binding sites of
histones are Lys rich, there have been many biophysical studies of DNA-cationic components as
model systems of DNA-histone interactions. However, electroneutral interpolyelectrolyte complexes
lead to precipitation and aggregation, and are therefore not suitable for quantitative analysis by
spectroscopy.
In order to establish experimental systems that mimic molecular environments inside cell nucleus,
here we utilize a comb-type copolymer of a polycation backbone and dextran side chains: poly(Llysine)-graft-dextran (PLL-g-Dex) (Fig.). PLL-g-Dex can form soluble interpolyelectrolyte complex with
DNA. The structure and thermodynamic stability of DNA oligonucleotides in the presence of PLL-gDex was studied. It was found that the parallel conformations involved in both DNA duplexes and the
DNA triplex were significantly and specifically stabilized by PLL-g-Dex, leading to higher stabilization
of the parallel DNA duplex than the antiparallel duplex under certain condition (3). These results
suggest that polycationic molecules are able to induce structural polymorphism of DNA
oligonucleotides, because of the conformation-selective stabilization effects.
References
1. S. Nakano, D. Miyoshi, N. Sugimoto, Chem.
Rev. 114, 2733 (2014).
2. H. Yaku, T. Murashima, D. Miyoshi, N.
Sugimoto, J. Phys. Chem. B 118, 2605 (2014).
3. D. Miyoshi et al. ChemMedChem 9, 2156
(2014).
Fig. Chemical structure of PLL-g-Dex
- 70 -
P-39
Design, Synthesis, and Evaluation of Phenyl-Piperazine-Triazine-Based α-Helix
Mimetics Targeting Protein-Protein Interactions
Heejo Moon, Woo-Sirl Lee, Misook Oh, and Hyun-Suk Lim*
Department of Chemistry, Pohang University of Science and Technology, Pohang 790-784, Korea
mhj0804@postech.ac.kr
Many cellular events occur via protein-protein interactions (PPIs) where protein secondary structures,
such as α-helix, often play an important role as a recognition motif. From the fact that α-helix accounts
more than 40% of protein secondary structure in nature, designing α-helical mimetics is of great
importance to modulate PPIs.
Strategies to mimic α-helical structure have been tried over past decades. One of the most remarkable
compounds is terphenyl structure in which three substituted phenyl rings exist in array. The low
solubility in water and difficulty in synthesis, however, were pointed out as the fatal problems that
prevent this scaffold from the further extensive application. Therefore, it has been a big issue among
chemical biologists to develop a new class of scaffold with better water solubility and synthetic
accessibility.
Here we describe the design and efficient solid-phase synthesis of phenyl-piperazine-triazine scaffold
as a novel class of α-helix mimetic small molecules. The scaffold showed good aqueous solubility.
Subsequent screening of a focused library of the designed molecules identified a potent and selective
inhibitor of MCL-1/BH3 interaction, demonstrating their ability to act as inhibitors of α-helix-mediated
PPIs. Consequently, our scaffold, along with simple synthetic route, will provide an excellent source of
PPI modulators.
P-40
Curvature Sensitive Membrane Perturbation by
Adenovirus-derived Amphypathic Peptides
Tomo Murayama, Sílvia Pujals, and Shiroh Futaki
Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan
murayama.tomo.44u@st.kyoto-u.ac.jp
The N-terminal amphipathic helical segment of Adenovirus internal protein VI (AdVpVI (33-55)) plays
an important role in the membrane disruption by inducing positive curvature [1]. AdVpVI is a capsid
protein that contributes to the viral infection through endosomal membranes. We prepared the peptide
corresponding to AdVpVI (33-55) (acetyl–GAFSWGSLWSGIKNFGSTVKNYG-amide) and the several
derivatives by Fmoc solid-phase synthesis. Their interaction modes with bilayers were examined by
using large unilamellar vesicles of different diameters and lipid compositions. In this presentation, we
discuss the selective leakage activity of AdVpVI (33-55) depending on the diameters of the vesicles, or
membrane curvature. The derivatives which have substituted hydrophobic faces showed non-selective
leakage activity and less interaction with acyl part of the lipid bilayer than the original sequence. This
result provides helpful information for designing membrane interacting peptides having novel functions.
Membrane curvature plays a crucial role in cellular functions, including membrane trafficking,
morphological changes, growth and division. Spatiotemporal delivery or detection would be possible
by targeting membrane curvature.
[1] Maier, O.; Wiethoff, M. C. Virology 2010, 402, 11-19.
- 71 -
P-41
Development of the Strategy for the Selective Chemical Modification in an Abasic Site
of Duplex DNA
Norihiro Sato, Gen-ichiro Tsuji, Kazumitsu Onizuka, Fumi Nagatsugi*
The Institute of Multidisciplinary Research for Advanced Materials (IMRAM), Tohoku University
nagatugi@tagen.tohoku.ac.jp
[Introduction] Recently, the molecular probes for the biomolecule have attracted growing attention,
and many reports have emerged on the development of new techniques for construction of molecular
probes. We have previously reported that the oligonucleotides containing 2-amino-6-vinylpurine
nucleotide (2-AVP, 1) exhibited efficient and selective crosslinking to thymine at the target site in
[1]
DNA. We have anticipated that 2-AVP can form a complex with thymine using the two hydrogen
bonds and reacted with thymine by forcing them into close proximity as shown in Fig. 1. Based on this
result, we have designed the strategy for a selective chemical modification of the nucleobase opposite
to an abasic site. It is expected that 2-AVP derivative (2) binds to the thymine with two hydrogen
bonds at the opposite site of the abasic site, in which should be a hydrophobic space, and reacts to
that thymine selectively. In this presentation, we report the synthesis of 2-AVP derivatives and the
evaluation of their reactivity.
Fig.1 Selective cross-linking
Fig.2 Selective chemical
modification in an abasic site
reaction with 2-AVP
Fig.3 The structure of 2-AVP probes for
modification in an abasic site
2a: R= (Arg)5
2b: R=
2c: R=
1: 2-AVP
[Results and Discussion] We have designed the new probes for reactions in an abasic site in duplex
DNA. These probes consist of 2-AVP as a reactive moiety and penta arginine (2a: RRRRR), acridine
(2b) or Hoechst (2c) as a binding moiety to duplex DNA with high affinity. The reactivities of
synthesized probes were evaluated using the duplex DNA containing an abasic site and full-matched
DNA. These probes (2a~2c) reacted to the thymine opposite to an abasic site with high selectivity and
did not react with full- matched DNA. These results indicated that the probes (2) might form hydrogen
bonding with thymine on the opposite to an abasic site and induce the alkylation at this site.
The highest reactivity by using of 2a was observed in an abasic site of duplex DNA, but 2a also
reacted with single strand DNA. We assumed that the undesired alkylation would be caused by
nonselective electrostatic interaction between penta arginine and phosphate group of DNA. AcridineAVP (2b) and Hoechst-AVP (2c) probes showed high selectivity to the duplex DNA containing an
abasic site and the reactivity of 2c was higher than that of 2b. We would like to present these results
in detail.
[1]
F. Nagatsugi and S. Imoto, Org. Biomol. Chem., 2011, 9, 257.
- 72 -
P-42
Solution structure of P450 from Bacillus megaterium by small-angle X-ray scattering
Ji-Hye Oh 1, §, Donghyuk Shin 1, §, Seungsu Han 1, Tae Yong Jee 1, Sei Young Lee 1, YangGyun Kim 2 and Sangho Lee 1*
1
Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746, Korea
2
Department of Chemistry, Sungkyunkwan University, Suwon 440-746, Korea
sangholee@skku.edu
Cytochrome P450s catalyze mono-oxygenation of organic compounds in vivo. P450 from Bacillus
megaterium (P450BM3) combines two essential components of a mono-oxygenase system,
cytochrome P450 and P450 reductase, into single polypeptide, thereby serving as a convenient
system to study biologically and pharmacologically important mono-oxygenation reactions. P450BM3
consists of three domains: heme, FMN and FAD domains. Molecular structures are available for
constituent domains of P450BM3. However, no structural information about the full-length P450BM3 in
solution is known. Here we report the solution structure of the full-length P450BM3 as well as those of
heme, heme-FMN and FAD domains using small-angle X-ray scattering (SAXS). Molecular envelopes
derived from SAXS data for heme, heme-FMN, and FAD domains were fitted well with the previously
reported crystal structures. Envelope for the full-length P450BM3 revealed that P450BM3 heme
domain is juxtaposed with FMN domain, consistent with a previously proposed model. Presence of
ionic liquids, known to deteriorate the enzymatic activity of P450BM3, did not seem to alter structural
change in solution. Taken together, our data establishes the solution structure of the full-length
P450BM3, which will serve as a structural template for further biochemical and physiological
characterizations.
P-43
Assembled, Mid-sized Agents that Modulate Protein-protein Interactions
Mai Tsubamoto3, Prakash Parvatkar1, Louvy Punzalan1, Jiashi Sun2, Nobuo Kato3, Motonari
Uesugi1, Junko Ohkanda1,*
1
2
Institute for Chemical Research, Kyoto University, Japan
Department of Pharmaceutical Sciences, University of South Florida, USA
3
ISIR, Osaka University, Japan
johkanda@scl.kyoto-u.ac.jp
In humans, more than 300,000 protein-protein interactions (PPIs) play critical roles in regulating
biological functions, and their dysregulation causes a large number of diseases. Synthetic agents that
control intracellular PPIs have gained much attention due to their large potential application in new
therapeutics. We focus on design of synthetic agents that recognize large protein surfaces, and
utilizing them to disrupt, stabilize, and detect PPIs that are responsible for regulating intracellular
signalling pathways. Our molecular design is based on the module assembly; small compounds are
designed for local protein surfaces, and are assembled to create mid-sized multivalent agents. For
example, the assembled enzyme inhibitors were found to disrupt PPIs between oncogenic K-Ras and
prenyltransferases at low M, resulting in suppression of K-Ras processing efficiently. This
assembling approach may open a way toward modulating intracellular PPIs by synthetic agents.
- 73 -
P-44
Artificial Glutamate Analogs as a Ligand for Neuronal Receptors
Masato Oikawa, Manami Chiba
Graduate School of Nanobioscience, Yokohama City University, Japan
moikawa@yokohama-cu.ac.jp
Ionotropic glutamate receptors (iGluRs) play a pivotal role in learning and memory by mediating the
majority of fast excitatory neurotransmission in the mammalian central nervous system (CNS). We
have previously studied construction of a molecular library of artificial glutamate analogs by diversityoriented synthesis (DOS) for discovery of selective ligands for iGluRs. Further synthetic study based
on the structure of the hit compounds successfully led us to identify IKM–159 as an antagonist
selective to (S)–2–amino–3–(3–hydroxy–5–methyl–4–isoxazolyl)propionic acid (AMPA) type iGluRs.
IKM–159 selectively inhibits GluA2- and GluA4-containing subtype of AMPA type iGluR. Furthermore,
the interactions of IKM–159 with GluA2 ligand-binding domain (LBD) have been clarified by
crystallographic study.
To improve the potency and selectivity of IKM–159 toward AMPA type
iGluRs, in the present work, we further study the structure–activity
relationships of IKM–159 analogs. Here, we report our synthetic studies
along this line of research based on the second generation diversity-oriented
synthesis of C-ring analogs of the artificial glutamate (IKM–159) starting
from 7–oxanorbornene prepared by tandem Ugi / Diels–Alder reaction of 2–
furaldehyde. Synthesis and the preliminary data on the biological activity will
be discussed.
P-45
Design of Highly Reactive Peptide Tag for Protein Labeling and Protein Functional
Analysis
Shigekazu Tabata1, Hirokazu Fuchida1, Munetsugu Kido1, Itaru Hamachi2, Ryuichi
Shigemoto3, Akio Ojida1
1
Graduate School of Pharmaceutical Sciences, Kyushu University, Japan, 2Department of Synthetic
Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Japan,
3
Institute of Science and Technology Austria, Austria.
ojida@phar.kyushu-u.ac.jp
Selective protein labeling with a small molecular probe is a powerful method for elucidating protein
functions in living cells. Unlik fluorescence proteins, labeling method using small molecular probes
allow the post-translational introduction of a variety of functional molecules into a target protein at any
appropriate time with controllable labeling efficiency. In this presentation, we report a new protein
labeling method that enables selective covalent modification of a tag-fused protein with small
molecular probes. We developed a complementary recognition pair composed of an oligo-aspartate
tag (CPYSAADAAADAAADAAAD) and multinuclear Zn(II) complexes (Zn(II)-DpaTyr) as a new tagprobe system for the selective protein labeling. We found that the peptide forms -helix upon binding
to the Zn(II) complex, by which the nucleophilicity of the cysteine residue largely increases to facilitate
the specific covalent labeling of the peptide tag. This labeling system was successfully applied to
protein functional analyses such as fluorescence bioimaging of protein under the live cell conditions.
- 74 -
P-46
Peptide microarrays for rapid analysis of peptide-RNA interactions
Sookil Park1, Jaeyoung Pai1 and Injae Shin1
1
National Creative Research Center for Biofunctional Molecules, Department of Chemistry, Yonsei
University, Seoul 120-749, Korea
injae@yonsei.ac.kr
RNA molecules play a pivotal role in most events taking place in cells. These biomolecules often have
a hairpin motif as one of their most frequently occurring secondary structural features. A rapid and
quantitative method to evaluate binding properties of hairpin RNAs to peptides using peptide
microarrays has been developed. The microarray technology was shown to be a powerful tool for
high-throughput analysis of RNA−peptide interactions by its application to profiling interactions
between 111 peptides and six hairpin RNAs. The peptide microarrays were also employed to measure
hundreds of dissociation constants (Kd) of RNA−peptide complexes. Our results reveal that both
hydrophobic and hydrophilic faces of amphiphilic peptides are likely involved in interactions with RNAs.
Furthermore, these results also show that most of the tested peptides bind hairpin RNAs with
submicromolar Kd values. One of the peptides identified by using this method was found to have good
inhibitory activity against TAR−Tat interactions in cells. Because of their great applicability to
evaluation of nearly all types of RNA−peptide interactions, peptide microarrays are expected to serve
as robust tools for rapid assessment of peptide−RNA interactions and development of peptide ligands
against RNA targets.
P-47
Ppa-iRGD-Q as a Facile Depot Forming Peptide for Effective Photodynamic Therapy
Sung Jun Park1, Hong-Jun Cho2, Sehoon Kim2, Yoon-Sik Lee1
1
School of Chemical and Biological Engineering, Seoul National University, South Korea
2
Korea Institute of Science and Technology, Seongbuk-Gu, South Korea
pitpind@naver.com
1
Controlled release of singlet oxygen ( O2) is of major interest in photodynamic therapy (PDT) against
cancer for effective treatment with minimal side effects. We report herein a photosensitizer
(pyropheophorbide a, Ppa) conjugated tumor targeting peptide iRGD, which has a cyclic form with a
quencher at the other end (Ppa-iRGD-Q). The Ppa-iRGD-Q peptide was designed to internalize into
cancer cells via protease cleavage. It could recognize cancer cells via integrin αvβ3 by the RGD motif,
and after protease cleavage, Ppa was internalized into the cells by the C-end rule. Peritumoral
injection of the Ppa-iRGD-Q formed a molecular depot, which remained stable at the tumor‟s tissue
environment allowing sustained release of the Ppa. The Ppa was well contained inside the tumor
allowing in vivo detection of the fluorescence signal of Ppa and also effective generation of singlet
oxygen. We suggest that Ppa-iRGD-Q peptide has the potentials for stable, controlled release of the
photosensitizer into the tumor cell for effective PDT.
- 75 -
P-48
Design and Synthesis of Molecules Targeting the Non-enzymatic Tau protein in cells
Tingting Chu1, Tian Qiu1, Yanmei Li1
1
Key Lab of Bioorganic Phosphorus Chemistry & Chemical Biology (Ministry of Education),
Department of Chemistry, Tsinghua University, Beijing, 100084, P.R. China.
liym@mail.tsinghua.edu.cn
Alzheimer's disease (AD) is one of the most common neurodegenerative diseases. It is characterized
by two histopathological hallmarks, extracellular deposits of β-amyloid and intracellular neurofibrillary
tangles (NFTs). The latter are composed of the abnormal aggregation of tau protein, which plays an
important role in stabilizing the microtubules. Recent studies demonstrate that Aβ requires tau protein
to induce the cytotoxicity, and tau protein reduction protects against Aβ-induced various defects.
However，tau protein is non-enzymatic which has no small molecule binding pockets. It is of great
challenge to make the non-enzymatic diseases-associated protein susceptible to small molecules
control equally. Herein, we design and synthesize a series of peptides which can target tau protein
and degrade it controllably. It is shown that partial degradation of tau induced by the peptides can
rescue the mislocalization of organelle caused by tau overexpression and decrease the toxicity of Aβ
mediated by high tau level.
P-49
Identification of Sub-Mitochondrion Proteome in Living Mammalian Cells By Using
Engineered Ascorbate Peroxidase (APEX)
Hyun Woo Rhee*
Department of Chemistry, Ulsan National Institute of Science and Technology (UNIST), Korea
rhee@unist.ac.kr
Mitochondrion is a very interesting cellular organelle. It has its own genome but most of its proteins
are encoded in nuclear genome and are delivered from cytosol. Mitochondrion generates most of
ATPs in mammalian cell and produces very important metabolites such as heme and phospholipids.
Also, mitochondrion works as a control tower of several important cellular pathways such as apoptosis.
All these distinctive biochemical functions are believed to happen systemically in separated space of
mitochondrion, however, its sub-organelle proteome has not been well studied because of lack of
purification method of sub-mitochondrion space. Recently, engineered peroxidase (APEX) has been
developed for generating short-lived biotin-phenoxyl radicals to label spatial-resolved endogenous
proteome of interest in living cells and we found our method using APEX mapped sub-mitochondrion
proteome successfully. In this session, recent mapping results of two sub-mitochondrial space,
mitochondrial matrix (Rhee and Zou, Science, 2013) & intermembrane space (Hung, Mol Cell, 2014)
will be shown and discussed. These two dataset are very complementary to each other and generates
interesting open questions in cellular biology.
- 76 -
P-50
Covalent labeling and real-time imaging of cytokine receptors on cell membrane using
DMAP-tethered cytokines
Shohei Uchinomiya1, Yosuke Takaoka2, Takahiro Hayashi3, Yoshiaki Fukuyama3, Itaru
Hamachi3
1
Department of Chemistry, National University of Singapore, Singapore
2
Department of Chemistry, Tohoku University, Japan
3
Department of Synthetic Chemistry and Biological Chemistry, Kyoto University, Japan
chmsu@nus.edu.sg
Cytokine receptors on cell membrane play key roles in many biological events such as cell growth,
proliferation and migration. It is also well known that these receptors are involved in the development
and growth of cancer. Therefore, deeper understanding of their functions is highly demanded for anticancer therapy. We describe a new method for covalent labeling of cytokine receptors with functional
probes on live cell surface. To achieve the covalent labeling of cytokine receptors, we developed the
cytokines tethered with 4-dimethylaminopyridine (DMAP), a catalyst for acyl transfer reaction. The
DMAP-tethered reactive cytokines (DMAP-cytokines) were readily constructed through the noncovalent interaction between oligo-histidine tag and Ni complex of nitrilotriacetic acid without any
purification. These DMAP-cytokines can be used as a catalyst the covalent labeling of epidermal
growth factor receptor (EGFR) and Neuropilin 1 transiently expressed on live HEK293T cells.
Moreover, the labeling of endogenous EGFR on live A431 cells and real-time imaging of its ligandinduced dynamics were also successfully achieved.
P-51
In vitro selection of macrocyclic peptides that trap the open state structure of
channelrhodopsin
Xiao Song1, Hideaki E. Kato2, Andrés D. Maturana3, Takayuki Katoh1, Yuki Goto1,
Osamu Nureki2, and Hiroaki Suga1
1
Department of Chemistry, Graduate School of Science, The University of Tokyo, Tokyo 113-0033,
Japan, 2Department of Biological Sciences, Graduate School of Science, The University of Tokyo,
Tokyo 113-0033, Japan, 3School of Agricultural Sciences, Graduate School of Bioagricultural
Sciences, Nagoya University, Nagoya 464-8610, Japan
hsuga@chem.s.u-tokyo.ac.jp
Channelrhodopsin is an ion channel derived from green algae. It mediates the transport of cations
across the plasma membrane upon stimulation by light. Regulation of the channel gating
(opening/closing) by light governs precise and rapid depolarization of the plasma membrane and，in
consequence, channelrhodopsin has been used in the field of optogenetics as a tool to control neural
activity, enabling researchers to understand the function/dysfunction of large neural circuits in various
1
animal models . However owing to an incomplete understanding of its gating mechanism, engineering
channelrhodopsin to improve functionality for application of optogenetics remains challenging.
In our laboratory, we had developed a platform technology, which referred to as the RaPID (Random
2
non-standard Peptide Integrated Discovery) system . This technology enables the rapid selection of
macrocyclic peptides with high-affinity to and high-selectivity for any desired target protein from a
13
highly diverse peptide library (~10 peptides). In this study, we have attempted to select macrocyclic
peptides that bind to channelrhodopsin using the RaPID system. The aim was to discover macrocyclic
peptides that stabilize the open state structure of channelrhodopsin, which could then be used as cocrystallization ligands for the determination of its open state structure. In this presentation, I will
discuss the methodology we used in the study as well as the properties of the selected peptides.
1. F. Lenno, et al., Annu. Rev. Neurosci., 2011, 34, 389-412.
2. Y. Yamagishi, et al., Chem. & Bio., 2011, 18, 1562-1570.
- 77 -
P-52
A Chemical Fluorescent Probe for the Detection of Aβ Oligomers.
Chai Lean Teoh1, Dongdong Su1, Srikanta Sahu1, Seong-Wook Yun1, Young-Tae Chang1,2
1
Laboratory Bioimaging Probe Development, Singapore Bioimaging Consortium, Singapore.
2
Department of Chemistry, National University of Singapore, Singapore.
teoh_chai_lean@sbic.a-star.edu.sg
Aggregation of amyloid β-peptide (Aβ) is implicated in the pathology of Alzheimer‟s disease (AD), with
the soluble, Aβ oligomeric species thought to be the critical pathological species. Identification and
characterization of intermediate species formed during the aggregation process is crucial to the
understanding of the mechanisms by which oligomeric species mediate neuronal toxicity and following
disease progression. Probing these species proved to be extremely challenging, as evident by the lack
of reliable sensors, due to their heterogeneous and transient nature. We describe here an oligomerspecific fluorescent chemical probe, BDO-1, developed through the use of the diversity-oriented
fluorescent library approach (DOFLA) and high-content, imaging-based screening. This probe enables
dynamic oligomer-sensing during fibrillogenesis and shows specific staining in AD brain sections.
P-53
Crystallographic studies of adenylate kinase from Streptococcus pneumoniae D39 in
new conformations
Trung Thanh Thach and Sangho Lee
Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746, Korea
sangholee@skku.edu
Adenylate kinases (AdKs; EC 2.7.3.4) play an essential role in metabolic monitoring and signaling
pathways in living cells by interconversion between ATP and AMP to two ADP molecules. We recently
determined the crystal structure of adenylate kinase from Streptococcus pneumoniae D39 (SpAdK) in
apo and inhibitor-bound forms. Here we report new crystal structures of the apo and inhibitor-bound
SpAdK diffracting to 1.96 Å and 1.65 Å resolution, respectively. The new apo SpAdK crystal belonged
to space group C2 with cell parameters a = 73.5 Å, b = 54.3 Å, c = 62.7 Å, β = 118.8° and revealed an
open conformation differing from the reported one with an r.m.s.d. value of 1.4 Å for Cα positions. The
inhibitor-bound SpAdK crystal belonged to space group P1 with cell parameters a = 53.9 Å, b =62.3 Å,
c = 63.0 Å, α = 101.9°, β =112.6°, γ =89.9°. Although the space group is similar to the reported one,
the crystal displayed different lengths of the unit cell and lattice contacts. These results demonstrate
that SpAdK can crystallize in different forms and that apo structure can adopt multiple conformations.
- 78 -
P-54
Fabrication of Self-Assembling Peptide Hydrogels from Short Designed Peptides for
Cell Culture
Hiroshi Tsutsumi, Hisakazu Mihara
Department of Bioengineering, Graduate School of Bioscience and Biotechnology, Tokyo Institute of
Technology, Japan
htsutsum@bio.titech.ac.jp
Designed self-assembling peptides have attracted much interest in the fabrication of nano- and microscaled biomaterials, because they can assemble into nano- and micro-structured materials such as
fibers and hydrogels in aqueous media. In addition, self-assembling peptides have advantages in
modulating molecular assembly and introducing functionality [1]. Recently, we have found that a
designed self-assembling peptide E1Y9 and E1Y9 derivatives with bio-active sequences to promote
cell-adhesion or cell-diferentiation co-assembled into highly networked nanofibers and hydrogels, and
these materials worked as a good scaffold to culture cells [2]. In this study, we have developed new
short self-assembling peptides based on different peptide design with E1Y9 peptides. Different selfassembling motifs are expected to be orthogonal pairs to fabricate a highly functionalized material
system.
New self-assembling peptides, u(FFiK)2 and u(FFiO)2, were designed as bolaamphiphilic peptides that
have a symmetric urea-linked diphenylalanine motif with zwitter ionic termini composed of Lys or Orn.
Circular dichroism and transmission electron microscope (TEM) observation revealed that both
peptides formed -sheet structures and self-assembled into nanofibers under neutral pH condition. In
addition, u(FFiK)2 and u(FFiO)2 formed self-supporting clear hydrogels at 0.2 and 0.5 wt%
concentration under physiological pH, respectively. Furthermore, u(FFiK) 2 formed hydrogels in cell
culture media and HeLa and HEK293 cells well adhered to the surface of u(FFiK) 2 hydrogels.
Therefore, the u(FFiK)2 hydrogel is a promising biomaterial scaffold to culture cells.
1. H. Tsutsumi, H. Mihara, Mol. BioSyst., 2013, 9, 609.
2. K. Fukunaga, H. Tsutsumi, H. Mihara, Biopolymers, 2013, 100, 731.
P-55
Luminescent Graphene Oxide/Peptide-Quencher Hybrids for Optical Detection of CellSecreted Proteases by Turn-on Response
Jin-Kyoung Yang,1 Seon-Yeong Kwak,1,2 Su-Ji Jeon,2 Hye-In Kim,2 Joonhyuk Yim,1 Homan
Kang,3 San Kyeong,1 Jong-Ho Kim,*2 and Yoon-Sik Lee*1,3
1
School of Chemical and Biological engineering, Seoul National University, Seoul, 151-742, Korea
2
Department of Chemical Engineering, Hanyang University, Ansan 426-791, Republic of Korea
3
Interdisciplinary Program in Nano-Science and Technology and Nano Systems Institute, Seoul
National University, Seoul 151-742, Republic of Korea, Fax: +82-(2)-880-1604
yslee@snu.ac.kr
Proteases are of great relevance to physiological and pathological processes, and thus, have been
used as biomarkers in the fields of biology, medicine and biotechnology. In this context, there have
been continuous interests to develop a simple, rapid and sensitive sensing tool for monitoring
protease activity over the past decades. Recently, detection strategy using fluorescence resonance
energy transfer (FRET) between substrate-dye and quencher has emerged as an alternative. In
particular, graphene oxide (GO) based sensing platform has attracted great attentions due to featured
properties of GO, such as biocompatibility, chemical affinity to biomolecules, and fluorescence
quenching ability. Here, we construct GO-peptide-quencher using inherent fluorescence of GO for
optical detection of protease activity. Prior to this, we examine the quenching efficiency of various
quencher molecules such as metallo protoporphyrin derivatives (MePPs) and QXL 570 and present their
quenching mechanism. This sensing system can detect protease activity by recovery of GO
fluorescence and successfully monitor cell-secreted MMP-2 in living cells, human hepatocytes HepG2.
- 79 -
P-56
Study on the Novel Chemical Probes to Analyze Protein-ligand Interactions
Kozo Yoneda, Yaping Hu, Masaki Kita, Hideo Kigoshi
Graduate School of Pure and Applied Sciences, University of Tsukuba, Japan
kozo1124@dmb.chem.tsukuba.ac.jp
Laser desorption ionization (LDI) of pyrene-containing compounds is observed without a matrix. So,
they are expected to be selectively detected by the label-assisted LDI-MS. Our aim is to develop a
new strategy to analyze the binding mode of bioactive ligands on target proteins by using their pyrene
derivatives (pyrene probes). After covalent binding between pyrene probes and target proteins,
enzymatic digestion and mass analysis may enable us to identify ligand-binding peptides without
further purification. In this study, we selected a marine macrolide aplyronine A and biotin as bioactive
ligands, and have synthesized their pyrene probes. Aplyronine A interacts with both actin and tubulin.
Design, photolabeling experimets of these pyrene probes, and the mass analysis of photoreacted
products will be presented.
P-57
Development of Highly Potent Macrocyclic Peptide Inhibitors Targeting
Cofactor-independent Phosphoglycerate Mutase
Hao Yu1, Patricia Dranchak2, James Inglese2, Hiroaki Suga1
1
Department of Chemistry, Graduate School of Science, the University of Tokyo, Tokyo, 113-0033,
Japan
2
National Center for Advancing Translational Sciences, National Institutes of Health, Rockville,
Maryland 20850, United States
hsuga@chem.s.u-tokyo.ac.jp
Treatments for parasitic nematode caused diseases, such as the lymphatic filariasis caused by Brugia
malayi, have not been well developed to date. The enzyme Cofactor-independent phosphoglycerate
mutase (iPGM) catalyzes the critical interconversion of 2-phosphoglycerate and 3-phosphoglycerate in
the glycolytic and gluconeogenic metabolic pathways that are essential for the growth of nematodes,
implicating iPGM as a potential drug target [1]. However, iPGM is considered to have low
“druggability”, and is thus not suitable for targeting with conventional small molecules [2]. Thus,
alternative approaches are required for the discovery of iPGM inhibitors, which may be useful in the
development of antifilarial therapies.
In this study, we employed a RaPID (Random non-standard Peptides Integrated Discovery [3])
technique to discover candidate macrocyclic peptide inhibitors of iPGM. Such an approach enables
the rapid selection of high affinity iPGM-binders from a genetic code reprogrammed peptide library
containing trillions of unique macrocyclic peptides. Several iPGM-binding molecules were identified
and in vitro inhibition assays showed these peptides to be strong inhibitors of iPGM.
In this presentation, we report the details of iPGM-binding macrocyclic peptides development,
including studies of bioactivity and chemical modification.
References
[1] Singh et al.(2013) Infectious Diseases of poverty, 2:5
[2] Crowther GJ, et.al (2014) PLoS Negl Trop Dis, 8(1): e2628.
[3] Y. Yamagishi et.al (2011) Chemistry & Biology, 18, 1562-1570
- 80 -
P-58
Mitochondria and Lysosome targetable Two-Photon Fluorescent Probes for
Hypochlorous Acid Detection and Imaging in Live Cells and Tissues
Lin Yuan1, Lu Wang1, Sung-Jin Park2, Keshari Bikram1, Juanjuan Peng2, Balasubramaniam
Sivaraman2, Young-Tae Chang1,2
1
Department of Chemistry, National University of Singapore, Singapore
Laboratory Bioimaging Probe Development, Singapore Bioimaging Consortium, Singapore
chmynl@nus.edu.sg
2
Hypochlorous acid (HOCl), as a highly potent oxidant, is a double-edged sword in biological system,
which serves as a “killer” for both invasions and hosts in the innate immune system. However, the
biological roles of HOCl in cellular signalling pathways and various diseases have not yet been fully
elucidated. One of the main reasons lies in the lack of effective chemical tools to measure the
generation and concentration of HOCl at subcellular levels. Herein, some new two-photon fluorescent
probes and their mitochondria (Mito-TP) and lysosome (Lyso-TP) targetable derivatives for HOCl
detection were reported. These probes exhibit fast response (within 1 s), excellent selectivity and
sensitivity (30 nM) toward HOCl. In cell imaging experiments, it is indicated for the first time that
endogenous HOCl can be generated simultaneously both in mitochondria and lysosome in
macrophages. In particular, these probes were further used to detect of HOCl in inflammation tissues
through two photon imaging. These probes could therefore serve as promising tools to help elucidate
the generation and biological functions of HOCl.
- 81 -
Index
First Name
Last Name
Country
Mikiko
Sodeoka
Japan
Jong Seog
Misao
Keigo
Satoshi
Ahn
Akishiba
Aoi
Arai
South Korea
Yoshinobu
Baba
Japan
Kyung-Hwa
Nils Anton
Matthew Wook
Baek
Berglund
Chang
South Korea
Singapore
Xing
Chen
Peng
Chen
China
China
Sang J.
Chung
South Korea
Ikuo
Fujii
Japan
Yukari
Fujimoto
Japan
Hiroto
Koichi
Koichi
Fujita
Fukase
Fukusima
Japan
Japan
Japan
Shiroh
Futaki
Japan
Zhihong
Sihyun
Yuichiro
Daewha
Chuen-Jiuan
Soonsil
Myungsu
Ji-ryang
Sangyong
Sang-yong
Nam-Young
Myeong-Gyun
Vasileios
Yuka
Guo
Ham
Hirayama
Hong
Huang
Hyun
Jang
Jang
Jon
Ju
Kang
Kang
Kargas
Kataoka
Hong Kong
Kazuya
Kikuchi
Japan
Yun Kyung
Young-O
Yo-Han
Gyuhee
Hak Joong
Jong Seung
Sung-Eun
Byeang Hyean
Kazushi
Masaki
Kim
Kim
Kim
Kim
Kim
Kim
Kim
Kim
Kinbara
Kita
South Korea
South Korea
South Korea
South Korea
South Korea
Japan
Japan
Singapore
Singapore
South Korea
Japan
South Korea
Taiwan
South Korea
South Korea
South Korea
South Korea
South Korea
Singapore
South Korea
Singapore
Japan
South Korea
South Korea
South Korea
Japan
Japan
Email
sodeoka@riken.jp
jsahn@kribb.re.kr
akishiba.misao.87m@st.kyoto-u.ac.jp
aoi@agr.nagoya-u.ac.jp
satoshi.arai.chem@gmail.com
babaymtt@apchem.nagoya-u.ac.jp
cleochem@nate.com
nilsab@bii.a-star.edu.sg
matthew_chang@nuhs.edu.sg
xingchen@pku.edu.cn
pengchen@pku.edu.cn
sjchung@dongguk.edu
fujii@b.s.osakafu-u.ac.jp
fujimotoy@chem.keio.ac.jp
t09301148@gunma-u.ac.jp
koichi@chem.sci.osaka-u.ac.jp
n135220b@yokohama-cu.ac.jp
futaki@scl.kyoto-u.ac.jp
chguo@ust.hk
sihyun@sookmyung.ac.kr
hirayama@dmb.chem.tsukuba.ac.jp
dw_hong@kaist.ac.kr
h291172@gate.sinica.edu.tw
hobois@snu.ac.kr
msj0910@yonsei.ac.kr
jiryang.jang@gmail.com
syjon@kaist.ac.kr
syju@yonsei.ac.kr
kang_nam_young@sbic.a-star.edu.sg
rukia123@unist.ac.kr
vasileiosk@bii.a-star.edu.sg
t10301041@gunma-u.ac.jp
kkikuchi@mls.eng.osaka-u.ac.jp
yunkyungkim@kist.re.kr
zerofive@snu.ac.kr
rladyhan@snu.ac.kr
khyhies77@naver.com
hakkim@korea.ac.kr
jongskim@korea.ac.kr
sungeun.kim86@gmail.com
bhkim@posetch.ac.kr
kinbara@tagen.tohoku.ac.jp
mkita@chem.tsukuba.ac.jp
- 82 -
Page
18
49
51
51
52
20
52,68
53
25
47
17
19
34
46
53,60
54
54
28,51,71
55
33
55
56,68
56,69
57
57
58
29
58
59
59
60
60
21
61
61
61,62
62
63
36
63
23
64
55,64,80
Masayasu
San
Yongan
Song-Yi
Jun-Seok
Kang Ju
Hojae
Su Seong
Hui
Kuwahara
Kyeong
Lee
Lee
Lee
LEE
Lee
Lee
Li
Japan
South Korea
Singapore
South Korea
South Korea
South Korea
South Korea
Yan Mei
Li
Yaw-Kuen
Li
Tsung-lin
Li
Xiang David
Li
Hyun-Suk
Hsien-Ya
Chun-Cheng
Lim
Lin
Lin
Chun-Hung Hans
Lin
Po-Chiao
Lin
Daisuke
Heejo
Tomo
Fumi
Jihye
Junko
Masato
Akio
Sookil
Sung Jun
Tian
Hyun-Woo
Uchinomiya
Taebo
Jae Kyung
Xiao
Miyoshi
Moon
Murayama
Nagatsugi
Oh
Ohkanda
Oikawa
Ojida
Park
Park
Qiu
Rhee
Shohei
Sim
Sohng
Song
Hiroaki
Suga
Japan
Naoki
Sugimoto
Japan
Chai Lean
TrungThanh
Teoh
Thach
Singapore
South Korea
Kouhei
Tsumoto
Japan
Hiroshi
Tsutsumi
Japan
Motonari
Uesugi
Japan
Jiangyun
Wang
Shih-Hsiung
Wu
China
Taiwna
Bengang
Jin-Kyoung
Kozo
Hao
Xing
Yang
Yoneda
Yu
Singapore
South Korea
China
Taiwna
Taiwna
China
South Korea
Taiwan
Taiwna
Taiwna
Taiwna
Japan
South Korea
Japan
Japan
South Korea
Japan
Japan
Japan
South Korea
South Korea
China
South Korea
Singapore
South Korea
South Korea
Japan
Singapore
South Korea
Japan
Japan
kuwahara@chem-bio.gunma-u.ac.jp
feanor05@snu.ac.kr
lee_yong_an@sbic.a-star.edu.sg
song@unist.ac.kr
junseoklee@chembiol.re.kr
dlrkdwn2tlfd@postech.ac.kr
hojaetolee@kaist.ac.kr
sslee@ibn.a-star.edu.sg
lee921@yonsei.ac.kr
liym@mail.tsinghua.edu.cn
ykl@cc.nctu.edu.tw
tlli@gate.sinica.edu.tw
xiangli@hku.hk
hslim@postech.ac.kr
pipis_lsy@hotmail.com
cclin66@mx.nthu.edu.tw
chunhung@gate.sinica.edu.tw
pclin@mail.nsysu.edu.tw
miyoshi@center.konan-u.ac.jp
mhj0804@postech.ac.kr
murayama.tomo.44u@st.kyoto-u.ac.jp
nagatugi@tagen.tohoku.ac.jp
some1005@nate.com
johkanda@scl.kyoto-u.ac.jp
moikawa@yokohama-cu.ac.jp
ojida@phar.kyushu-u.ac.jp
chemskpark@yonsei.ac.kr
pitpind@naver.com
bekidl@gmail.com
rhee@unist.ac.kr
chmsu@nus.edu.sg
tbsim@kist.re.kr
sohng@sunmoon.ac.kr
x-song@chem.s.u-tokyo.ac.jp
hsuga@chem.s.u-tokyo.ac.jp
sugimoto@konan-u.ac.jp
chai.teoh@gmail.com
tttrung@skku.edu
tsumoto@k.u-tokyo.ac.jp
htsutsum@bio.titech.ac.jp
uesugi@scl.kyoto-u.ac.jp
jwang@ibp.ac.cn
Shwu@gate.sinica.edu.tw
bengang@ntu.edu.sg
yjk0627@snu.ac.kr
kozo1124@dmb.chem.tsukuba.ac.jp
yuhaocool@gmail.com
- 83 -
53,60,65
65,79
66
66
67
67
56,68
31
68
30
48
40
22
27,67,71
69
37
44,68
43
70
71
71
72
73
73
54,74
74
75
75
76
59,66,76
77
42
45
77
35,77,79
24,70
78
78
38
79
41,55,73
26
32
39
79
55,80
80
Lin
Yuan
Singapore
CHMYNL@nus.edu.sg
- 84 -
81