title: Intramolecular binding of a proximal PPII helix to an SH3 domain in the fusion protein SH3Hck : PPIIhGAP.
abstract: SH3 domains are a conserved feature of many nonreceptor protein tyrosine kinases, such as Hck, and often function in substrate recruitment and regulation of kinase activity. SH3 domains modulate kinase activity by binding to polyproline helices (PPII helix) either intramolecularly or in target proteins. The preponderance of bimolecular and distal interactions between SH3 domains and PPII helices led us to investigate whether proximal placement of a PPII helix relative to an SH3 domain would result in tight, intramolecular binding. We have fused the PPII helix region of human GAP to the C-terminus of Hck SH3 and expressed the recombinant fusion protein in Escherichia coli. The fusion protein, SH3Hck : PPIIhGAP, folded spontaneously into a structure in which the PPII helix was bound intramolecularly to the hydrophobic crevice of the SH3 domain. The SH3Hck : PPIIhGAP fusion protein is useful for investigating SH3: PPII helix interactions, for studying concepts in protein folding and design, and may represent a protein structural motif that is widely distributed in nature.
authors.affiliation: Department of Biochemistry, Wake Forest University School of Medicine, Winston-Salem, NC 27157, USA. bgmeiner@wfubmc.edu
authors.forename: W H
authors.initials: WH
authors.lastname: Gmeiner
authors.forename: I
authors.initials: I
authors.lastname: Xu
authors.forename: D A
authors.initials: DA
authors.lastname: Horita
authors.forename: T E
authors.initials: TE
authors.lastname: Smithgall
authors.forename: J R
authors.initials: JR
authors.lastname: Engen
authors.forename: D L
authors.initials: DL
authors.lastname: Smith
authors.forename: R A
authors.initials: RA
authors.lastname: Byrd
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D004058:Diffusion; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D006859:Hydrogen / Q000378:metabolism; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D020703:ras GTPase-Activating Proteins / Q000737:chemistry*; D018909:src Homology Domains
chemical_list: D008024:Ligands; D010455:Peptides; D011993:Recombinant Fusion Proteins; D020703:ras GTPase-Activating Proteins; C011083:polyproline; D006859:Hydrogen