A NOVEL MAMMALIAN RAS GTPASE-ACTIVATING PROTEIN WHICH HAS PHOSPHOLIPID-BINDING AND BTK HOMOLOGY REGIONS

We have previously purified a novel GTPase-activating protein (GAP) fo r Ras which is immunologically distinct from the known Ras GAPs, p120G AP and neurofibromin (M. Maekawa, S. Nakamura, and S. Hattori, J. Biol . Chem. 268:22948-22952, 1993). On the basis of the partial amino acid sequence, we have obtained a cDNA which encodes the novel Ras GAP. Th e predicted protein consists of 847 amino acids whose calculated molec ular mass, 96,369 Da, is close to the apparent molecular mass of the n ovel Ras GAP, 100 kDa. The amino acid sequence shows a high degree of similarity to the entire sequence of the Drosophila melanogaster Gap1 gene. When the catalytic domain of the novel GAP was compared with tha t of Drosophila Gap1, p120GAP, and neurofibromin, the highest degree o f similarity was again observed with Gap1. Thus, we designated this ge ne Gap1(m), a mammalian counterpart of the Drosophila Gap1 gene. Expre ssion of Gap1(m) was relatively high in brain, placenta, and kidney ti ssues, and it was expressed at low levels in other tissues. A recombin ant protein consisting of glutathione-S-transferase and the GAP-relate d domain of Gap1(m) stimulated GTPase of normal Ras but not that of Ra s having valine at the 12th residue. Expression of the same region in Saccharomyces cerevisiae suppressed the ira2(-) phenotype. In addition to the GAP catalytic domain, Gap1(m) has two domains with sequence cl osely related to those of the phospholipid-binding domain of synaptota gmin and a region with similarity to the unique domain of Btk tyrosine kinase. These results clearly show that Gap1(m) is a novel Ras GAP mo lecule of mammalian cells.