Abstract
A general assay method for nucleotide pyrophosphorylases has been investigated. The principle of the method is based on the measurement of consumption rate of 5-phos-phoribosylpyrophosphate (PRPP) during the enzyme reaction. In the method, an enzyme preparation for sample was incubated in a reaction mixture containing a purine or pyri-midine base and PRPP for a certain time, and the amounts of PRPP before and after the reaction were determined. The amount of PRPP was determined by an enzymatic method using orotidine-5'-monophosphate (5'-OMP) pyrophosphorylase and 5'-OMP de-carboxylase. Nucleotide pyrophosphorylase activity corresponding to each purine or pyri-midine base was determined from the amount of PRPP consumed per unit time.
The present method is generally applicable for determining activities of any kind of nucleotide pyrophosphorylases, and does not need any tedious separation procedure in all cases. Therefore, comparing with the conventional assay methods for nucleotide pyrophos-phorylase activities, this method can be said to be much simpler and reliable. As an application of the present method, activities of several nucleotide pyrophosphorylases in Micrococcus glutamicus have been determined.
