in silico discovery of inhibitors using structure-based approaches jasmita gill structural and...
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In silico discovery of inhibitors using
structure-based approaches
Jasmita Gill
Structural and Computational Biology Group,
ICGEB, New Delhi
Nov 2005
Target protein 3D structure
Find an inhibitor
Molecular modeling
In silico screening
Computational Techniques
Computational approach
In silico screening
Structure based virtual screening
docking methods to fit putative ligands into 3D structure of target receptor
Structure-based inhibitor discovery
3D structure of target protein Public drug-like in silico libraries
In silico screening
Short listed hits provided for testing in biological assays
Binding site (s) identification
Post-scoring and analysis of results
Literature, Visual analysis
FlexX
Cscore, Visual analysis, Unity
VendorsProtein Data Bank
Sybyl® – Molecular modelling suite
Tools and Techniques
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
Input
OutputEnergetically best ranked ligand placements in target site (s)
Each placement has variable conformations
Target protein with pre-defined active site (s)
and
Ligands with designated base fragment (s)
Thomas Lengauer et. al, J Mol. Bio. 1996
- Multiple conformations determined by torsion angles of
acyclic single bonds in the ligands
- Low energy conformation of the complex is the goal
Considerations in FlexX
Receptor target protein rigid
Ligand Conformational Flexibility
Modeling protein-ligand interactions
Interaction geometries
protein ligand
Interactions types
H-acceptor H-donor
Metal acceptor Metal
Aromatic-ring-atom,
Methyl, amide
Aromatic-ring-center
Main scoring criteria
Free energy of binding of protein-ligand
Consensus scoring ‘Cscore’
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
Final output of screening:
Ranking based on free energy of binding of protein-ligand complex
Visual
Mathematical
Binding sites to which compounds docked
Conformations
H-bonding interactions
Hydrophobic interactions
Van Der Waals attractions
Cscore
Screening results
Analysis
Application to Pfg27
Binding sites of interest on Pfg27
• Two RNA binding sites per dimer
• Four SH3 binding sites per dimer
• A dimer interface
From literature
• Revealed a deep cavity on a unique surface
Visual/computational analysis
RNA binding site
SH3 binding site (N)
Dimer interface
RNA binding site
Deep cavity
Colour coding
Basic
Acidic
Non-polar
Polar
Deep cavity
Depth
Surface
Deepest cavity in Pfg27
Cavities in the dimer interface
Cavities
SH3 binding site
Cavity
Cavities in the SH3 binding site (N)
Cavities in the RNA binding site
Multiple cavities of different depths
NCI-diversity set: 1820 compounds
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
Docking patterns on Pfg27
Visual analysis of top 200
• Best binding energies observed:
from -44.363 KJ/mol to –24.056 KJ/mol
• 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)
• 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
Analysis of top 200 compounds
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