[Isolation and radiometric determination of rat liver ATP: thiamine diphosphate phosphotransferase activity]

A radioassay was developed for estimation of the thiamin diphosphate kinase activity (EC 2.7.4.15); 14C-TDP was used as a substrate. After termination of the reaction (formation of TTP), the products obtained were separated using ion exchange chromatography on SP-Sephadex C-25. The method developed was very sensitive and enabled to estimate the enzymatic activity in tissue homogenates. TDP-kinase was isolated from rat liver with a 70-fold purification. Dependence of the reaction rate on pH value, concentrations of the enzyme and thiamin were studied.