An enzymatic method for the measurement of GTP and GDP in biological samples

✍ Scribed by F.A.M. de Azeredo; G.K. Feussner; W.D. Lust; J.V. Passonneau

• Publisher: Elsevier Science
• Year: 1979
• Tongue: English
• Weight: 598 KB
• Volume: 95
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(79)90764-4

✦ Synopsis

A method is described for the analysis of GTP/GDP, or GDP alone, in a coupled enzymatic assay. Samples are pretreated with creatine kinase in the presence of phosphocreatine to remove the ADP present. Succinyl CoA synthetase is used to convert GTP to GDP in the presence of succinate and coenzyme A. Endogenous GDP and that formed from GTP are measured in a second step with pyruvate kinase and lactate dehydrogenase in the presence of excess NADH. The sensitivity of the assay is in the nanomole range. A further modification is described, where the final product NAD+ is cycled in yet another enzyme system, extending the sensitivity to the picomole range. Sample analyses are given for a variety of tissues using 7 mg of sample and for 1-pg freeze-dried sections from the cerebellum of mouse brain ' The U. S. Government's right to retain a nonexclusive royalty-free license in and to the copyright covering this paper, for governmental purposes, is acknowledged.

* Recipient of a postdoctoral fellowship from Con-