Ha-ras proteins exhibit GTPase activity: point mutations that activate Ha-ras gene products result in decreased GTPase activity.
Several ras genes have been expressed at high levels in Escherichia coli and the resultant ras proteins were shown to be functional with respect to their well-known specific, high-affinity, GDP/GTP binding. We were able to detect a weak GTPase activity associated with the purified proteins. The normal cellular ras protein (p21N) exhibits approximately equal to 10 times higher GTPase activity than the "activated" proteins. Even though the turnover rate of the reaction is very low (0.02 mol of GTP hydrolyzed per mol of p21N protein per minute), the reaction appears to be catalytic; one molecule of p21N hydrolyzes more than one molecule of GTP. The GTPase and the GDP binding activities both have been recovered from a Mr 23,000 protein eluted following NaDodSO4/polyacrylamide gel electrophoresis, suggesting that these two activities are associated with the same protein. Mg2+ ions and dithiothreitol are required for GTPase activity and the optimal pH is between 7 and 8. Guanidine X HCl, which is required for solubilizing bacterially expressed ras protein, is strongly inhibitory to GTPase activity at concentrations higher than 0.5 M.
Manne
V
V
Bekesi
E
E
Kung
H F
HF
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D019791:Guanidine; D006146:Guanidines; D006153:Guanosine Diphosphate / Q000378:metabolism; D006251:Harvey murine sarcoma virus / Q000235:genetics; D008274:Magnesium / Q000494:pharmacology; D009154:Mutation; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D009053:Sarcoma Viruses, Murine / Q000235:genetics; D013997:Time Factors
D001426:Bacterial Proteins; D006146:Guanidines; D006153:Guanosine Diphosphate; D010744:Phosphoric Monoester Hydrolases; D020558:GTP Phosphohydrolases; D008274:Magnesium; D019791:Guanidine; D004229:Dithiothreitol