Structural biology should be
computable!
• Protein structures determined by amino acid
sequences
• Protein structures and complexes correspond
to global free energy minima
• Fundamental test of understanding and huge
practical relevance
Model of energetics of
inter and intramolecular
interactions
Prediction
(Given
Sequence,
Optimize
Structure)
Ab initio structure
prediction
ROSETTA
Protein Structure
Protein-protein docking Protein-protein
interactions
Design
(Given
Structure,
Optimize
Sequence)
Protein design
Interface design
Model of macromolecular
interactions
• Removal of single methyl groups can
destabilize proteins --> jigsaw puzzle-like
packing crucial
• Buried polar atoms almost always hydrogen
bonded --> treat hydrogen bonding as
accurately as possible
• Exposed charge substitutions generally have
little effect --> damp long range elctrostatics
• Focus on short range interactions!
Conformational
sampling
Random Start
Low-Resolution
Monte Carlo Search
(integrate out
sidechain degrees of
freedom)
High-Resolution
Refinement with
full atomic detail
105
Jeff Gray (Hopkins),
Ora Furman (Hebrew University), Chu Wang
Select
lowest
energy
models
Predictions
Docking Low-Resolution Search
•
•
•
Monte Carlo Search
Rigid body translations and
rotations
Residue-scale interaction
potentials
Protein representation: backbone
atoms + average centroids
O
O
N
...
N
N
O
O
N
O
O
N
...
Docking Protocol
(Target 12: cohesin-dockerin; unbound-bound)
2. Refinement
Energy
1. Initial Search
RMSD to arbitrary starting structure
(Å)
RMSD to starting structure of refinement
Side Chain Flexibility
Target 12
Cohesin-Dockerin
dockerin
 0.46Å interface rmsd
 87% native contacts
 6% wrong contacts
Ora Furman,
Chu Wang
red,orange– xray
blue – model;
green – unbound
cohesin
Details of T12 Interface
dockerin
R53
S45
D39
L22
Y74
N37
L83
E86
cohesin
red,orange– xray
blue - model
Accurate Side Chain Modeling
Target 15
immunity protein Dcolicin D tRNase
colicin
 0.23Å interface rmsd
Science 310, 638-642
immunity protein
red,orange– xray
blue - model
Details of T15 Interface
colicin
H611
K610
K607
K608
E56
E68
D61
E59
immunity protein
red,orange– xray
blue - model
Modeling Backbone Movement
Target 20
HemK-RF1
 2.34Å
interface rmsd
 36% native
contacts
RF1
HemK
red,orange– xray
blue – model;
green – unbound
Loop with methylated Gln
Chu Wang
QuickTime™ and a
YUV420 codec decompressor
are needed to see this picture.
CASP6 T0198: PhoU domain repeat
Phil Bradley
Model 2: 4A over 210 rsds
(Model 1: 3.94 over 198)
CASP6 T0212
Model 2: 3.97 over 109 rsds
(Model 1: 4.0 over 104)
QuickTime™ and a
DV/DVCPRO - NTSC decompressor
are needed to see this picture.
T0281 ab initio prediction (1.59Å)
Phil Bradley
1r69
Science 309, 1868-1871
1ubq
2REB
Boinc.bakerlab.org/rosetta
David Kim
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
High resolution ab initio structure prediction from
single sequences by enhanced diversity “barcode”
directed sampling
Outreach!
High Resolution Refinement of CASP target 199 - remote homology model
Bin Qian
Calculations performed on SDSC teragrid clusters
High Resolution NMR Model Refinement
Vatson Raman
Disulfide Bond
Formation Protein
Blue - X-ray structure Green - NMR models Red - Rosetta models
Computing Structural Biology
• Free energy function reasonable =>
Computing simple protein structures and
interactions now appears to be within reach
• Implications for structural genomics?
• More cpu power => more accurate
predictions for larger proteins
• For larger complexes, experimental data
essential (low resolution electron density!).
• Symmetry helps!
Modeling accuracy also illustrated by structures
of designed proteins
Top7 X-ray structure has correct topology.
Backbone RMSD to design only 1.2Å!!
C-a Backbone Overlay
Red : X-ray structure
Blue : Design model
Brian Kuhlman, Gautam Dantas;
Science 302 1364-8
Design of novel H bond network
Q51
Design
Q51
X-ray
Y35
Y35
Y35
Q169
Q169
Q180
G177
interface
Lukasz Joachimiak
Q180
G177
G177
Design of new protein
functions
• Design of new protein-protein interactions
• Design of enzymes catalyzing novel
chemical reactions
• Design of new transcription factor and
endonuclease specificities
• Design of HIV vaccine
HIV vaccine design
• Present HIV coat protein epitopes locked into
conformation observed in complexes with
neutralizing antibodies using designed
scaffolds
• Preliminary results: designed proteins fold
and bind neutralizing antibodies (5nM
affinity). One design confirmed
crystallographically.
Bill Schief in collaboration with Peter Kwong
Computational design of non-HIV immunogens
to elicit broadly-neutralizing antibodies
Bill Schief
Crystal structure of Mab 2F5
in complex with its HIV epitope
Model of non-HIV
scaffold-epitope (red)
Redesign of
DNA cleavage
specificity of MsoI
homing endonuclease
using ROSETTA
Justin Ashworth,
Jim Havranek
Nature in press
WT-WT
Design-WT
WT-Design
Design-Design
Specific DNA cleavage by designed nuclease
1
½
¼
1/2n
1/29
-
wild-type
design
5uM
nuclease
Cleavage
targets
wild-type I-Mso
-
wild-type
design
Design
Acknowledgements
Protein structure prediction
• Phil Bradley (MIT)
• Rhiju Das
• Lars Marlstrom
• Bin Qian
• Vatson Raman
Protein-protein docking
• Ora Furman (Hebrew
University)
• Chu Wang
• Jeff Gray (Johns
Hopkins)
Design
• Brian Kuhlman (UNC)
• Gautam Dantas
• Justin Ashworth
• Jim Havranek
Robetta.bakerlab.org
prediction and design server:
David Kim (domain parsing,
boinc) and Dylan Chivian
Rosetta software freely
available for academic use
Boinc.bakerlab.org/rosetta