SubCav - Tool for subpocket comparison and alignment

Dr. Tuomo KalliokoskiLead Discovery Center GmbH, Dortmund, Germany

Work conducted atNovartis Institutes for Biomedical Research, Basel, Switzerland

Kalliokoski T, Olsson TSG, Vulpetti A. J. Chem. Inf. Model. 2013, 53, 131-141.

Protein Databank (PDB) is growing

• Number of searchable structures 1972-Mar 2013

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How many fragments are there?

8 million unique chemical structures

2 million lead-like structures

400,000 Rule-Of-Three compliant structures

Zuegg and Cooper. Drug-Likeness and Increased Hydrophobicity of Commercially Available Compound Libraries for Drug Screening. Curr Top Med Chem 2012, 12, 1500-1513.

Bridging “Structural”-Space and “Fragment”-Space

Fragment chemical space is too large for experimentalFragment-Based Drug Design (FBDD)

The need to develop tools for FBDD to take

advantage of PDB!

The information content of PDB is increasing

Binding site similarity

“The availability of such data provides a basis for the identification of bioisosteres that are target specific. The resulting bioisosteres might be expected to provide more reliable information when modifying an existing lead compound than do existing approaches, which are based either on empirical measures of inter-substituent similarity or on non-target specific crystallographic data.”

Kennewell EA, Willett P, Ducrot P, Luttmann C. Identification of target-specific bioisosteric fragments from ligand–protein crystallographic data. J Comput Aided Mol Des 2006, 20, 385-394.

Subpockets and fragments

*Degen J, Wegscheid-Gerlach C, Zaliani A, Rarey M. On the art of compiling and using ’drug-like’ chemical fragment spaces. ChemMedChem 2008, 3, 1503–1507.

BRICS*

SubCav

• Tool for subpocket similarity searching and alignment

• Based on pharmacophoric fingerprints with geometric hashing-inspired alignment

• Source code available via anna.vulpetti@novartis.com

Fingerprint descriptorSubCav atom type PDB atom types

Acceptors with sp2 character (π-acceptor) (A=)

ALA.O ARG.O ASN.O ASN.OD1 ASP.O ASP.OD1 ASP.OD2 CYS.O GLN.O GLN.OE1 GLU.O GLU.OE1 GLU.OE2 GLY.O HIS.O ILE.O LEU.O LYS.O MET.O PHE.O PRO.O SER.O THR.O TRP.O TYR.O VAL.O

α-carbon (CA) ALA.CA ARG.CA ASN.CA ASP.CA CYS.CA GLN.CA GLU.CA GLY.CA HIS.CA ILE.CA LEU.CA LYS.CA MET.CA PHE.CA PRO.CA SER.CA THR.CA TRP.CA TYR.CA VAL.CA

Donor (D) LYS.NZ

Donors with sp2 character (π-donor) (D=)

ALA.N ARG.N ARG.NE ARG.NH1 ARG.NH2 ASN.N ASN.ND2 ASP.N CYS.N GLN.N GLN.NE2 GLU.N GLY.N HIS.N ILE.N LEU.N LYS.N MET.N PHE.N SER.N THR.N TRP.N TRP.NE1 TYR.N VAL.N

Hydrophobe (H) ALA.CB ARG.CB ARG.CD ARG.CG ASN.CB ASP.CB CYS.CB CYS.SG GLN.CB GLN.CG GLU.CB GLU.CG HIS.CB HIS.CG ILE.CB ILE.CD1 ILE.CG1 ILE.CG2 LEU.CB LEU.CD1 LEU.CD2 LEU.CG LYS.CB LYS.CD LYS.CE LYS.CG MET.CB MET.CE MET.CG MET.SD PHE.CB PRO.CB PRO.CD PRO.CG SER.CB THR.CB THR.CG2 TRP.CB TYR.CB VAL.CB VAL.CG1 VAL.CG2

π-hydrophobe (H=) HIS.CD2 HIS.CE1 PHE.CD1 PHE.CD2 PHE.CE1 PHE.CE2 PHE.CG PHE.CZ TRP.CD1 TRP.CD2 TRP.CE2 TRP.CE3 TRP.CG TRP.CH2 TRP.CZ2 TRP.CZ3 TYR.CD1 TYR.CD2 TYR.CE1 TYR.CE2 TYR.CG TYR.CZ

neutral donor & acceptor (P)

HIS.ND1 HIS.NE2 SER.OG THR.OG1 TYR.OH

Ignored PRO.N and all HETATM

Bin Range (Å)1 2.1-4.52 4.5-6.33 6.3-8.04 8.0-10.0

3.4Å=1

9.3Å=4

6.0Å=2

A=

D=

CA

Alignment algorithm

Implementation details

Validation study

• Align pairwise all similar subpockets in PSMDB* (non-redundant subset of PDB)

• 3,268,620 pairs from 3,886 PDBs with 17,044 subpockets with 332 different fragments

• Two alignment methods:– Fragment-based alignment– SubCav-based alignment

* Wallach I, Lilien R. The Protein–Small-Molecule Database (PSMDB), A Non-Redundant Structural Resource for the Analysis of Protein-Ligand Binding, Bioinformatics 2009, 25, 615-620.

When are two subpockets similar?• Two subpockets are similar if both after

alignment have– Root-Median-Square-Deviation (RMSD) of

fragments found in subpockets is less than 1.5 Å– Enough matched features*

*Matched feature=if two features from the two subpockets are within 1 Å distance

RMSD = 1.00Overlap = 0.79

Very rarely subpockets with same fragments are geometrically similar...

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Fragment-based OKSubCav- based OKBoth OKNot matched

SubCav finds 73%-85% of fragment-based (plus something else!)

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Three structures of thrombin aligned. The query (magenta) fragment-aligned (green) vs. SubCav aligned (cyan)

Bioisosteric replacement example

ACP

Heat Shock Protein 90 (HSP 90)

Bioisosteric replacement example

Escherichia coli DNA gyrase B(sequence similarity 30%)

Bioisosteric replacement example

Escherichia coli DNA gyrase B(sequence similarity 30%)

Adenine -> pyrazole?

Bioisosteric replacement example

HSP90 inhibitor

Analysis of Histone Methyl-Transferase Binding Sites

S-adenosylmethionine (SAM) or S-adenosyl-l-homocysteine (SAH) Fragmented in three: adenine, ribose, and tail fragments

Pairwise SubCav-alignment and hierarchical clustering based on Overlap

Analysis of Histone Methyl-Transferase Binding Sites

The clustering of the cofactor binding site by subpockets around each specific fragment revealed different levels of local similarity within the selected proteins set.

Analysis of Histone Methyl-Transferase Binding Sites

The clustering of the cofactor binding site by subpockets around each specific fragment revealed different levels of local similarity within the selected proteins set.

Analysis of Histone Methyl-Transferase Binding Sites

A B C D

Take home message

Subpocket analysis can provide ideas in CADD

Acknowledgements

• Novartis Institutes for Biomedical Research:– Dr. Anna Vulpetti (mentor & co-author)– Education office (Presidential Postdoctoral

Fellowship)• Cambridge Crystallographic Data Centre:

– Dr. Tjelvar Olsson (mentor & co-author)• Chemical Computing Group:

– Dr. Guido Kirsten (idea for alignment protocol)