Identification of amino acid residues of Ras protein that are essential for signal-transducing activity but not for enhancement of GTPase activity by GAP.
To determine the amino acid residues required for the signal-transducing activity of the human c-Ha-Ras protein, we introduced point mutations at residues 45-54 near the 'effector region' (residues 32-40). We transfected PC12 cells with these mutant genes and also micro-injected the mutant proteins, bound with an unhydrolyzable GTP analog, into PC12 cells. Both procedures showed that Val45----Glu and Gly48----Cys mutations impaired the ability of the Ras protein to induce morphological change of PC12 cells. These mutations did not affect the guanine nucleotide-binding activity or GTPase activity in the absence or presence of bovine GTPase-activating protein (GAP). Therefore, the Val45 and Gly48 residues should be included by definition in the effector region responsible for the signal transduction, while only a subset of the effector-region residues is required for enhancement of the GTPase activity by GAP.
Fujita-Yoshigaki
J
J
Department of Biophysics and Biochemistry, Faculty of Science, University of Tokyo, Japan.
Shirouzu
M
M
Koide
H
H
Nishimura
S
S
Yokoyama
S
S
D000310:Adrenal Gland Neoplasms; D000596:Amino Acids / Q000737:chemistry*; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D011905:Genes, ras; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010673:Pheochromocytoma; D011506:Proteins / Q000494:pharmacology*; D016283:Proto-Oncogene Proteins p21(ras) / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014162:Transfection; D014407:Tumor Cells, Cultured; D020703:ras GTPase-Activating Proteins
D000596:Amino Acids; D020690:GTPase-Activating Proteins; D011506:Proteins; D020703:ras GTPase-Activating Proteins; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D016283:Proto-Oncogene Proteins p21(ras)