Intrinsic GTPase activity distinguishes normal and oncogenic ras p21 molecules.
- 1 September 1984
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 81 (18), 5704-5708
- https://doi.org/10.1073/pnas.81.18.5704
Abstract
The 21-kilodalton protein (p21) encoded by normal cellular Harvey-ras was expressed in Escherichia coli as a fusion protein by using the pUC8 vector and was purified to > 95% homogeneity by ion-exchange chromatography and gel filtration. The purified protein molecules possess intrinsic GTPase activity on the basis of the following criteria: elution of the GTPase activity with p21 GDP-binding activity in 2 different chromatography systems, parallel thermal inactivation of GTPase activity and p21 GTP-binding activity, and immunoprecipitation of the GTPase activity with monoclonal antibodies to p21. At 37.degree. C, the rate of GTP hydrolysis by the purified normal p21 assayed in solution was 5.3-6.6 mmol/min/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/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. 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-GTP complex.This publication has 31 references indexed in Scilit:
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