Multiprotein assemblies, protein-protein interfaces and modulation of cell activity

Transcription

1 Multiprotein assemblies, protein-protein interfaces and modulation of cell activity Implications for drug discovery Defining 3D Structures of Multiprotein Assemblies Protein-protein Interfaces Tools for Modulating Protein Interfaces Cell Surface Receptor Clusters Intracellular Signalling Complex DNA-Repair Signalling Assembly Tom Blundell Cambridge ProteinProteinProtein Protein Interaction Interfacial Regions Credo: Protein Ligand Interactions Chemical Tool? Drug Candidate?

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3 Achieving Specificity in Regulatory and Signalling Systems Receptor Binding through Preformed Globular Structure Preformed globular structure Insulin hexamer in storage in pancreas Preformed globular structures Insulin monomer binds receptor Discontinuous epitopes Preformed but adaptive globular structures

4 Achieving Specificity in Regulatory and Signalling Systems Receptor Binding through Concerted Folding & Binding Concerted Folding & Binding Glucagon trimer in storage un pancreas Glucagon folds on binding receptor One partners becomes structured on assembly Continuous epitopes

5 Achieving Specificity in Regulatory and Signalling Systems Receptor Activation and Signal Transduction In the 1990s. u Binary complexes are likely to occur opportunistically u They will lead to high signal to noise unless they are very high affinity u But they have to be reversible to retain regulatory sensitivity u Multiprotein complexes can form cooperatively from weak binary complexes, so ensuring high signal to noise!

6 X-ray structure Pellegrini, Blundell et al., Nature, 406, Nano-spray Mass Spectrometry Nicholas J. Harmer, Leopold L. Ilag, Barbara Mulloy, Carol V. Robinson,Tom Blundell

7 Achieving Specificity in Regulatory and Signalling Systems u Weak dimers followed by co-operative formation of multiprotein systems u Binding at Hotspot/Foothold through Anchor Residue followed by Cooperative Folding and Binding So ensuring high signal to noise!

8 The Human Proteome Multiprotein Systems in Space and Time

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10 The Human Proteome Multiprotein Systems in Space and Time Why are multi-protein systems important How do they exhibit co-operativity? How do we define structures of multi-protein systems? In space and time? How should we organise information multi-protein systems?

11 CREDO database of protein interactions represents contacts as structural interaction fingerprints, sequence-to-structure mapping molecular shape descriptors with Ultrafast Shape Recognition (USR), fragmentation of ligands in PDB, identification of approved drugs. completely scriptable through application programming interface.

12 Targeting Protein-protein Interactions in Drug Discovery Why are multi-protein systems important How do they exhibit co-operativity? How do we define structures of multi-protein systems? In space and time? How do we organise information multi-protein systems? How do we extend information multi-protein systems?

13 Extending Knowledge of the Structural Interactome Database Organization and Modeling Pipeline

14 Structural Interactome FGF-stimulated MAPK pathway displayed as a network of interaction Proteins involved in FGF pathway (pale blue ellipses) are expanded (dotted lines) into constituent SCOP domains (pale orange squares 23 Binary interactions have been extracted from GLORIA17 based on these SCOP domains. The pathway is from KEGG pathway database Lee et al. (2009) Molecular BioSystems doi: /B906402H

15 Targeting Protein-protein Interactions in Drug Discovery Why are multi-protein systems important How do they exhibit co-operativity? How do we define structures of multi-protein systems? In space and time? How do we organise information multi-protein systems? How do we extend information multi-protein systems? What do protein-protein interfaces look like? Flat, uninteresting and large?

16 Surveying the structural landscape of proteinprotein interfaces Kawabata T (2010). Detection of multiscale pockets on protein surfaces using mathematical morphology.proteins; 78(5): Harry Jubb Harry Jubb

17 Targeting Protein-protein Interactions in Drug Discovery

18 Targeting the Interactome TIMBAL a hand-curated database holding current small molecules inhibiting multi-protein complexes. Give insights into type of molecules favoured by protein interfaces Used to analyse protein-protein modulators which tend to be big lipophilic molecules with few hydrogen bond features. Access at TIMBAL

19 Nature does more polar!!!! open closed medium

20 Targeting Protein-protein Interactions in Drug Discovery

21 Targeting Multiprotein Systems Exploiting Allostery Modulating Conformation Classical Allosteric Modulators

22 Pockets which could bind SSR located in conserved regions allosteric inhibition?

23 Targeting Multiprotein Systems Exploiting Allostery Modulating Conformation Classical Allosteric Modulators Modulating Location through Protein-Protein Interactions in Multiprotein Systems Scaffolds Adaptors Regulatory Assemblies

24 Established in December Founded by: Harren Jhoti, Glaxo Wellcome Tom Blundell, University of Cambridge Chris Abell, University of Cambridge Harren Jhoti CEO > 80 employees Protein Technology Structural Biology Original funding $500K Chemo-informatics Medicinal Chemistry Three people in the Pharmacology Biochem & Chem Labs Clinical Medicine Two above the local shopping centre Tom Blundell Non-executive Board Member & Chair of SAB

25 Fragments usually have millimolar affinities Neutral hydrogen bonds (cdk2) Lipophilic (p38) Neutral hydrogen bonds, lipophilic (p38) Lipophilic (thrombin) Hartshorn et al, J Med Chem 2005,403

26 The power of the cancer research and drug discovery clusters in Cambridge UK has prompted Japanese pharma giant Otsuka to buy Science Park medical technology pioneer Astex in an $886 million deal. 2012: Eight Candidate Drugs in Clinical Trials

27 Targeting Multiprotein Systems Exploiting Allostery Modulating Conformation Classical Allosteric Modulators Modulating Location through Protein-Protein Interactions in Multiprotein Systems Scaffold Adaptors Regulatory Assemblies

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29 Fragment hits u 1338 fragments screened using thermal shift u 81 induced shifts > 0.8 C selected for further analysis u Validation by NMR, ITC and X-ray KD = 540 µm KD = 1000 µm KD = 730 µm KD = 430 µm KD = 600 µm Slide devised by Marko Hyvonen

30 Increasing the potency K D = 2.0 mm K D = 0.6 mm K D = 0.4 mm K D =3 µm K D = 1 µm (K D = 80 nm) Thermal shi+ screen 1 st comp. NMR screen 2 nd comp. NMR screen Fragments + Pep:des Chemical elabora:on Ongoing op:misa:on Dec 2007 Aug 2008 Oct 2008 Mar 2009 Jul 2010 Oct 2011