SELECTION AND RADIOMETRIC METHOD OF DETERMINING THE ACTIVITY OF ATP: THIAMINEDIPHOSPHATEPHOSPHOTRANSFERASE FROM MITOCHONDRIA OF THE PIG’S BRAIN TISSUE

Introduction. The purpose and biological pathways of regulation of level of thiamine triphosphate ester in the cellremain unclear. This is due to the absence of reliable methodological basis for the assessment of the activity of the enzyme of the biosynthesis of this metabolite - ATP: thiaminediphosphatephosphotransferase (EС 2.7.4.15). Objective. To develop an adequate method of separation of ingredients of transferase reaction, to isolate a partially purified drug of the enzyme. Material and methods. As substratewas usedan exogenous 14С– thiaminediphosphate, that was obtained by us.Thehaving beenresulted product – thiamine triphosphate ester was separated by ion-exchange chromatography. Results. From the mitochondrial fraction of the pig's brain was isolated a partially purified transferase preparation with a yield of 14.2%. It was found that the maximum enzymatic activity is manifested at pH 7.0 - 8.5, the concentrations of thiamine diphosphate is 2.5 - 3.0∙10-5 M. Conclusions. The proposed isotope method has sufficient sensitivity and allows fixing ATP: thiamine diphosphate phosphotransferase activity in subcellular fractions of the brain.