PDBsum entry 4b3c

Hydrolase

PDB id: 4b3c

Contents

Protein chain

224 a.a.

Ligands

PO4

5H1

Waters ×153

PDB id:

4b3c

Name:

Hydrolase

Title:

Humanised monomeric rada in complex with 5-hydroxy indole

Structure:

DNA repair and recombination protein rada. Chain: a. Fragment: atpase, residues 108-287,300-349. Engineered: yes. Mutation: yes

Source:

Pyrococcus furiosus. Organism_taxid: 2261. Expressed in: escherichia coli. Expression_system_taxid: 469008. Expression_system_variant: containing pubs520 plasmid.

Resolution:

1.90Å R-factor: 0.210 R-free: 0.261

Authors:

D.E.Scott,M.T.Ehebauer,T.Pukala,M.Marsh,T.L.Blundell, A.R.Venkitaraman,C.Abell,M.Hyvonen

Key ref:

D.E.Scott et al. (2013). Using a fragment-based approach to target protein-protein interactions. Chembiochem, 14, 332-342. PubMed id: 23344974

Date:

23-Jul-12 Release date: 06-Feb-13

Protein chain

O74036  (RADA_PYRFU) -  DNA repair and recombination protein RadA from Pyrococcus furiosus (strain ATCC 43587 / DSM 3638 / JCM 8422 / Vc1)

Seq: 349 a.a.

Struc: 224 a.a.*

* PDB and UniProt seqs differ at 5 residue positions (black crosses)

Chembiochem 14:332-342 (2013)

PubMed id: 23344974

Using a fragment-based approach to target protein-protein interactions.

D.E.Scott, M.T.Ehebauer, T.Pukala, M.Marsh, T.L.Blundell, A.R.Venkitaraman, C.Abell, M.Hyvönen.

ABSTRACT

The ability to identify inhibitors of protein-protein interactions represents a major challenge in modern drug discovery and in the development of tools for chemical biology. In recent years, fragment-based approaches have emerged as a new methodology in drug discovery; however, few examples of small molecules that are active against chemotherapeutic targets have been published. Herein, we describe the fragment-based approach of targeting the interaction between the tumour suppressor BRCA2 and the recombination enzyme RAD51; it makes use of a screening pipeline of biophysical techniques that we expect to be more generally applicable to similar targets. Disruption of this interaction in vivo is hypothesised to give rise to cellular hypersensitivity to radiation and genotoxic drugs. We have used protein engineering to create a monomeric form of RAD51 by humanising a thermostable archaeal orthologue, RadA, and used this protein for fragment screening. The initial fragment hits were thoroughly validated biophysically by isothermal titration calorimetry (ITC) and NMR techniques and observed by X-ray crystallography to bind in a shallow surface pocket that is occupied in the native complex by the side chain of a phenylalanine from the conserved FxxA interaction motif found in BRCA2. This represents the first report of fragments or any small molecule binding at this protein-protein interaction site.