title: Intrinsic GTPase activity distinguishes normal and oncogenic ras p21 molecules.
abstract: The 21-kilodalton protein (p21) encoded by normal cellular Harvey-ras has been expressed in Escherichia coli as a fusion protein by using the pUC8 vector and has been purified to greater than 95% homogeneity by ion-exchange chromatography and gel filtration. The purified protein molecules possess intrinsic GTPase activity on the basis of the following criteria: (i) elution of the GTPase activity with p21 GDP-binding activity in two different chromatography systems, (ii) parallel thermal inactivation of GTPase activity and p21 GTP-binding activity, and (iii) immunoprecipitation of the GTPase activity with monoclonal antibodies to p21. At 37 degrees C, the rate of GTP hydrolysis by the purified normal p21 assayed in solution was 5.3-6.6 mmol/min per mol of p21. The rate of GTP hydrolysis by a form of p21 [Val12] encoded by a human oncogene was significantly lower (1.4-1.9 mmol/min per mol of p21). The presence of a threonine phosphate acceptor site at residue 59 also decreased p21 GTPase activity. For regulatory proteins that use GTP as part of their biochemical mechanism, the hydrolysis of GTP to GDP reverses the biological activity of the respective proteins. The observation that oncogenic forms of p21 lose GTPase activity suggests that GTP hydrolysis may be a biochemical event that inactivates the growth-promoting effects of a p21 X GTP complex.
mesh_terms: D000818:Animals; D004926:Escherichia coli / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D005822:Genetic Vectors; D051379:Mice; D009154:Mutation; D009363:Neoplasm Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D009857:Oncogenes; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D010957:Plasmids; D011485:Protein Binding; D016283:Proto-Oncogene Proteins p21(ras)
chemical_list: D009363:Neoplasm Proteins; D010761:Phosphorus Radioisotopes; D010744:Phosphoric Monoester Hydrolases; D020558:GTP Phosphohydrolases; D016283:Proto-Oncogene Proteins p21(ras)