Regulatory role of creatine phosphate in the phosphofructokinase system

Abstract

Phosphofructokinase from rabbit skeletal muscle is an allosteric enzyme as a key regulatory site in the glycolysis sequence. The regulatory characteristics of phsophofructokinase were studied by using coupled assay methods. Phosphofructokinase activity was inhibited by high concentrations of substrate ATP, and the degree of ATP inhibition was dependent on the concentrations of the second substrate fructose 6-phosphate, i.e. the lower level of fructose 6-phosphate was, the higher degree of ATP inhibition appeared. Initial rate profile as a function of fructose 6-phosphate was sigmoidal, but at the very low concentrations of ATP, hyperbolic nature was also observed. Under the high concentration of ATP and/or low concentration of fructose 6-phosphate, the enzyme activity was affected by the presence of creatine phosphate, i.e. inhibition of enzyme activity and stimulation of ATP inhibition were observed by the increasing concentrations of creatine phosphate. The extent of substrate inhibition by ATP and also the sigmoidicity as a function of fructose 6-phosphate were increased remarkably by the presence of creatine phosphate. ITP could act as substrate but could not cause substrate inhibition, in contrast with ATP. When ITP was used as substrate, creatine phosphate did not have any effect on the enzyme activity. It is believed that creatine phosphate act as a modulator in the phosphofructokinase system, and it seems that the regulatory role of creatine phosphate is very closely related to the ATP inhibition. Based on these results from kinetic experiments and some chemical modifications, the regulatory role of creatine phosphate in phosphofructokinase reaction was suggested in relation to their possible role in glycolytic regulation.