title: Grb7-SH2 domain dimerisation is affected by a single point mutation.
abstract: Growth factor receptor bound protein 7 (Grb7) is an adaptor protein that is co-overexpressed and forms a tight complex with the ErbB2 receptor in a number of breast tumours and breast cancer cell lines. The interaction of Grb7 with the ErbB2 receptor is mediated via its Src homology 2 (SH2) domain. Whilst most SH2 domains exist as monomers, recently reported studies have suggested that the Grb7-SH2 domain exists as a homodimer. The self-association properties of the Grb7-SH2 domain were therefore studied using sedimentation equilibrium ultracentrifugation. Analysis of the data demonstrated that the Grb7-SH2 domain is dimeric with a dissociation constant of approximately 11 muM. We also demonstrate, using size-exclusion chromatography, that mutation of phenylalanine 511 to an arginine produces a monomeric form of the Grb7-SH2 domain. This mutation represents the first step in the engineering of a Grb7-SH2 domain with good solution properties for further biophysical and structural investigation.
authors.affiliation: School of Biomedical and Chemical Sciences, University of Western Australia, 35 Stirling Highway, Crawley, Perth, WA 6009, Australia.
authors.forename: Corrine J
authors.initials: CJ
authors.lastname: Porter
authors.forename: Matthew C J
authors.initials: MC
authors.lastname: Wilce
authors.forename: Joel P
authors.initials: JP
authors.lastname: Mackay
authors.forename: Peter
authors.initials: P
authors.lastname: Leedman
authors.forename: Jackie A
authors.initials: JA
authors.lastname: Wilce
mesh_terms: D001120:Arginine / Q000737:chemistry; D001703:Biophysics / Q000379:methods; D045744:Cell Line, Tumor; D002845:Chromatography; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015233:Models, Statistical; D008968:Molecular Conformation; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010649:Phenylalanine / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D014461:Ultracentrifugation; D018909:src Homology Domains
chemical_list: D010649:Phenylalanine; D001120:Arginine