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.

Pubmed ID: 9607145 RIS Download