Transcript Jasmita_presen_nov3

In silico discovery of inhibitors using
structure-based approaches
Jasmita Gill
Structural and Computational Biology Group,
ICGEB, New Delhi
Nov 2005
Computational approach
Target protein 3D structure
Find an inhibitor
Computational Techniques
Molecular modeling
In silico screening
In silico screening
Structure based virtual screening
docking methods to fit putative ligands into 3D structure of target receptor
Structure-based inhibitor discovery
Protein Data Bank
3D structure of target protein
Vendors
Public drug-like in silico libraries
Binding site (s) identification
In silico screening
Literature, Visual analysis
FlexX
Post-scoring and analysis of results
Short listed hits provided for testing in biological assays
Cscore,
Visual analysis,
Unity
Tools and Techniques
Sybyl® – Molecular modelling suite
Analysis of molecular surfaces of proteins
Preparation of target protein and ligand(s) for screening
Screening utility -- FlexX
Post-scoring -- Cscore
Data Mining -- Unity
Public in silico chemical compound libraries used
FlexX – an overview
Target protein with pre-defined active site (s)
Input
and
Ligands with designated base fragment (s)
Output
Energetically best ranked ligand placements in target site (s)
Each placement has variable conformations
Thomas Lengauer et. al, J Mol. Bio. 1996
Considerations in FlexX
Receptor target protein rigid
Ligand Conformational Flexibility
-
Multiple conformations determined by torsion angles of
acyclic single bonds in the ligands
-
Low energy conformation of the complex is the goal
Modeling protein-ligand interactions
Interactions types
Interaction geometries
H-acceptor
H-donor
Metal acceptor
Metal
Aromatic-ringatom,
Aromatic-ringcenter
Methyl, amide
Main scoring criteria
Free energy of binding of protein-ligand
Consensus scoring ‘Cscore’
protein
ligand
Public drug-like in silico libraries
•
A database of structures of small molecule compounds
•
Most libraries are free to download
•
Lead-like properties
•
Available for purchase
Name
No. of Compounds
NCI Diversity set
NCI Open Collection
1990
~200,000
Maybridge
~95,000
Specs
~202,000
Peakdale
~20,000
In silico Screening
 Preparation of the target protein structure
Templates for charged, neutral, non-polar residues
Charges
Hydrogens

Preparation of ligand structure
Charges
Hydrogens
Filtering was done based on Lipinski’s rule of 5
Mw < 500 daltons (relaxed, <=900)
H-bond acceptors < 10
H-bond donors < 5
ClogP (solubility indicator) < 5
 Definition of binding site (s) : whole protein in case of Pfg27
Screening results
Final output of screening:
Ranking based on free energy of binding of protein-ligand complex
Analysis
Mathematical
Cscore
Visual
Binding sites to which compounds docked
Conformations
H-bonding interactions
Hydrophobic interactions
Van Der Waals attractions
Application to Pfg27
Binding sites of interest on Pfg27
From literature
• Two RNA binding sites per dimer
• Four SH3 binding sites per dimer
• A dimer interface
Visual/computational analysis
• Revealed a deep cavity on a unique surface
RNA binding site
RNA binding site
SH3
binding
site (N)
Deep
cavity
Dimer interface
Colour coding
Basic
Acidic
Non-polar
Polar
Depth
Deepest cavity in Pfg27
Deep cavity
Surface
Cavities in the dimer interface
Cavities
Cavities in the SH3 binding site (N)
Cavity
SH3 binding site
Cavities in the RNA binding site
Multiple cavities of
different depths
NCI-diversity set: 1820 compounds
Docking patterns on Pfg27
Visual analysis of top 200
30% in the RNA binding site
30% in the dimer interface
20% in deep cavity
10% in SH3 binding site (N)
10% on other sites
Analysis of top 200 compounds
• Best binding energies
observed:
from -44.363 KJ/mol to –24.056 KJ/mol
• Cscore
3 to 5 (a good score is 4-5)
Score of 3 – 37 compounds
Score of 4 – 43 compounds
Score of 5 – 48 compounds
• Chemical composition
Most hits had an electronegative character:
N, O-, SO3-, Cl-, F-, Br• CLogP: –3.59 to 1 (-4 to 4 range is acceptable for solubility)
Dockings in the RNA binding site
Most compounds interact with
Arg70, Arg74, Arg78, Arg80 and Val71
Dockings in the deep cavity
Most compounds interact with
Ser107, Lys112 and Ile122
Dockings at the dimer interface
Most compounds interact with
Asp40, Arg36, Glu134, Arg131,
Phe43, Leu126, Trp127
Dockings in SH3 binding sites
Most hits interact with Arg34