Comparative study of p120 GTPase-activating protein and its point mutant in the pleckstrin homology domain.
GTPase-activating protein (GAP) enhances the intrinsic GTPase activity of cellular Ras. In addition to two Src homology 2 (SH2) domains and one Src homology 3 (SH3) domain, it contains a pleckstrin homology (PH) domain. The wild-type or point mutant in the PH domain of p120 GAP (W568A) was expressed by using the baculovirus/Sf9 cell system. Direct effects of the G protein beta gamma subunit (G beta gamma) and several sphingolipids and the effects of phosphorylation by c-Src on the GTPase-stimulating activity of these GAPs on Ras were examined by using immunoprecipitates of these GAPs. The activities of neither of these GAPs were affected by the addition of G beta gamma, although the W568A mutant bound less to G beta gamma compared with the wild type. Several sphingolipids had no effect on the activity of these GAPs. Only in the W568A mutant was GTPase-stimulating activity reduced by tyrosine phosphorylation by c-Src.
Nakata
H
H
Department of Pharmacology, National Defense Medical College, Saitama, Japan.
Watanabe
Y
Y
D001798:Blood Proteins / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000502:physiology; D020690:GTPase-Activating Proteins; D006801:Humans; D015688:Oncogene Protein pp60(v-src) / Q000378:metabolism; D010750:Phosphoproteins; D010766:Phosphorylation; D017354:Point Mutation / Q000502:physiology*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D017386:Sequence Homology, Amino Acid; D013107:Sphingolipids / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism; D020703:ras GTPase-Activating Proteins; D018631:ras Proteins / Q000378:metabolism*
D001798:Blood Proteins; D020690:GTPase-Activating Proteins; D010750:Phosphoproteins; D011506:Proteins; D011993:Recombinant Fusion Proteins; D013107:Sphingolipids; C039077:platelet protein P47; D020703:ras GTPase-Activating Proteins; D014443:Tyrosine; D015688:Oncogene Protein pp60(v-src); D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D018631:ras Proteins