title: Crystal structure of the ARF-GAP domain and ankyrin repeats of PYK2-associated protein beta.
abstract: ADP ribosylation factors (ARFs), which are members of the Ras superfamily of GTP-binding proteins, are critical components of vesicular trafficking pathways in eukaryotes. Like Ras, ARFs are active in their GTP-bound form, and their duration of activity is controlled by GTPase-activating proteins (GAPs), which assist ARFs in hydrolyzing GTP to GDP. PAPbeta, a protein that binds to and is phosphorylated by the non-receptor tyrosine kinase PYK2, contains several modular signaling domains including a pleckstrin homology domain, an SH3 domain, ankyrin repeats and an ARF-GAP domain. Sequences of ARF-GAP domains show no recognizable similarity to those of other GAPs, and contain a characteristic Cys-X(2)-Cys-X(16-17)-Cys-X(2)-Cys motif. The crystal structure of the PAPbeta ARF-GAP domain and the C-terminal ankyrin repeats has been determined at 2.1 A resolution. The ARF-GAP domain comprises a central three-stranded beta-sheet flanked by five alpha-helices, with a Zn(2+) ion coordinated by the four cysteines of the cysteine-rich motif. Four ankyrin repeats are also present, the first two of which form an extensive interface with the ARF-GAP domain. An invariant arginine and several nearby hydrophobic residues are solvent exposed and are predicted to be the site of interaction with ARFs. Site-directed mutagenesis of these residues confirms their importance in ARF-GAP activity.
authors.affiliation: Department of Pharmacology, Skirball Institute of Biomolecular Medicine, New York University Medical School, New York, NY 10016, USA.
authors.forename: V
authors.initials: V
authors.lastname: Mandiyan
authors.forename: J
authors.initials: J
authors.lastname: Andreev
authors.forename: J
authors.initials: J
authors.lastname: Schlessinger
authors.forename: S R
authors.initials: SR
authors.lastname: Hubbard
mesh_terms: D020727:ADP-Ribosylation Factors / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D017487:Ankyrins / Q000737:chemistry* / Q000378:metabolism; D002384:Catalysis; D003196:Computer Graphics; D018360:Crystallography, X-Ray / Q000379:methods; D051418:Focal Adhesion Kinase 2; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D018909:src Homology Domains
chemical_list: D017487:Ankyrins; D011994:Recombinant Proteins; D011505:Protein-Tyrosine Kinases; D051418:Focal Adhesion Kinase 2; D020727:ADP-Ribosylation Factors