A method for continuous monitoring of phosphorylase b activity during glycogen degradation and synthesis

Abstract

Phosphorylase b (1,4-α-glucaniorthophosphate glucosyltransferase, EC. 2.4.1.1) catalyzes the glycogen synthesis and degradation reaction. It is the latter which is the physiological function of the enzyme. Its action may be described by a random order bi-bi kinetic mechanism. The rate-limiting step is an interconversion between ternary complexes (1). Activity determination can be carried out in the direction of glycogen synthesis by measuring the amount of inorganic phosphate (P j) released during the reaction (2). This method requires termination of the enzyme action after an appropriate time interval and only yields information about the total amount of the product without giving details of the time course of the process. In the reverse direction, the glycogen degradation can be followed in a more complicated and indirect way by coupled reactions of additional enzymes (3). The present communication describes a convenient method for continuous monitoring of the reaction in both directions with a fluorescence spectrometer.