NMR Solution Structure of HP0827 (O25501_HELPY)
from Helicobacter pylori: Model of the possible RNA
Binding Site
In-gyun Lee, Sun-Bok Jang, Chao Ma, Ji-Yoon Lee, Ji-Hun Kim, Sung Jean Park
and Bong-Jin Lee
Research Institute of Pharmaceutical Sciences, College of Pharmacy, Seoul
National University, San 56-1, Shillim-Dong, Kwanak-Gu, Seoul 151-742, Korea
Corresponding author: Research Institute of Pharmaceutical Sciences, College of
Pharmacy, Seoul National University
Phone: +82-2-880-7869; Fax: +82-2-872-3632; E-mail: lbj@nmr.snu.ac.kr.
Abstract
The HP0827 protein is a 82 residue protein identified as a putative ss-DNA Binding Protein
12RNP2 Precursor from Helicobacter pylori. Here, we have determined 3D structure of HP0827
using NMR. It has a ferredoxin-like fold, β1-α1-β2-β3-α2-β4 (α; α helix and β; β sheet) and RNP
motifs which are thought to be important in RNA binding. By using structural homologues search and
analyzing electrostatic potential of surface, we could compared HP0827 with other RNA binding
proteins (sex-lethal, Tia-1, U1A) to predict RNA binding sites of HP0827. We could predict that β
sheets of HP0827, especially β1 and β3, are primary region for RNA binding. Consequently, similar to
other RNA binding proteins, RNP motifs (Y5, F45, F47), positively charged and hydrophobic
regions (K32, R37, K40, K41, K43, R70, R73) are proposed as a putative RNA binding sites. In
addition, differences in amino acids composition of RNP motifs, N, C-terminal residues, loop region
fold and the orientation of α1 helix with other RRM proteins could give specific biological functions
to HP0827. Finally, the study on natural RNA target is also important to completely understand the
biological function of HP0827.
Physicochemical
Properties
of
Pyrazinamidase
from
Mycobacterium
tuberculosis
Su-Jin Cho, Woo-sung Son, Won-Je Kim, Yong-Jin Kim, Se-Won Suh, Takahisa Ikegami, Hideo
Akutsu & Bong-Jin Lee
physical Pharmacy, Department of Manufacturing Pharmacy
The Graduate School, Seoul National University
Physicochemical properties of Pyrazinamidase (PncA), which converts PZA, prodrug for TB,
to active form, were investigated. Structural information was obtained using circular
dichroism (CD), nuclear magnetic resonance (NMR), homology modelling, and molecular
dynamics (MD), and enzymatic activity was monitored by isothermal titration calorimetry in
acidic pH and neutral pH. Three-dimensional structure with active site, which was
surrounded by aromatic residues, was acquired and flexibility of structure was predicted.
PncA from M. tuberculosis appears to be more effective and stable hydrolytic enzyme in
neutral pH than in acidic pH and enzymatic reaction was little affected by zinc metal ion. The
results of study implicate that aromatic residues in active site lead to unique feature of M.
tuberculosis PncA with regard to hydrolysis and zinc coordination and this feature is related
to PZA-resistance mechanism in TB.
Characterization of a novel toxin-antitoxin module HP0894 and
HP0895, encoded by Helicobacter pylori chromosome
Yeon-Jin Yang, Hong-Man Kim1, Kyung-Doo Han1, Atsushi Matsuura1, 1, Hee-Chul
Ahn2, Sung-Jean Park3, Bong-Jin Lee1
1
College of Pharmacy, Seoul National University, Seoul, South Korea
2
Advanced Analysis Center, Korea Institute of Science and Technology, Seoul, South
Korea
3
Department of Pharmacology, Gachon university of Medicine and Science, Incheon,
South Korea
Some bacterial plasmids and chromosomes have toxin-antitoxin(TA) systems, which
specify two proteins; one works as a toxin, and the other as an antitoxin. Toxins are
neutralized by forming tight complexes with antitoxins. Previously we reported the
solution structure of the conserved hypothetical protein HP0894 from Helicobacter
pylori. It was found that it has significant structural similarities with that of archaeal
RelE toxin of Pyrococcus horikoshii. Herein we report that HP0894 and HP0895
of Helicobacter pylori are homologous to a toxin-antitoxin system. HP0894 is a YafQhomologous toxin with intrinsic RNase activity, and HP0895 is its antitoxin. Using NMR
titration experiment, we demonstrated that HP0894 binds to HP0895 strongly with a
stoichiometry ratio of 1:1, and the interaction between HP0894 and HP0895 occurs
mainly through N-terminal region of HP0894 and unstructured C-terminal region of
HP0895. Furthermore, HP0894 recognizes mRNA substrates with mainly three clusters
of residues(site 8KS9,site 50KGKWK54 and E58, and site L78,R80 and 84HSELF88). It
cleaves -U:A- or -C:A- sequences predominantly, and the recognition of these
specific sequences occurs mainly through the residues K52,W53,S85,E86 and K87 of
HP0894.
Keywords: Helicobacter pylori; Nuclear Magnedtic Resonance (NMR); HP0894; HP0895 ;
Toxin_Antitoxin ; RNase