Pyruvate kinase isozymes in adult tissue and eggs of Rana pipiens. II. Physical and kinetic studies of purified skeletal and heart muscle pyruvate kinases

Abstract

1. 1. Pyruvate kinase (ATP:phosphotransferase, EC 2.7.1.40) from skeletal muscle of Rana pipiens has been purified to homogeneity as judged by sedimentation velocity studies, immunoelectrophoresis, double diffusion experiments and acrylamide-gel electrophoresis. The enzyme has a molecular weight of approx. 220 000 as estimated by gel-filtration studies and a subunit molecular weight of approx. 55 000 as measured by acrylamide-gel electrophoresis in the presence of sodium dodecyl sulfate. 2. 2. The properties of the frog skeletal and partially purified cardiac pyruvate kinase have been compared. Both enzymes have the same molecular weight as estimated by gel-filtration experiments and the cardiac enzyme cross-reacts with the antibody to the skeletal muscle pyruvate kinase. However, the cardiac enzyme exhibits different chromatographic and kinetic properties. Unlike the muscle enzyme, heart pyruvate kinase is retained on DEAE-Sephadex, displays temperature sensitivity, is inhibited by l-alanine and dl-phenylalanine and is activated by fructose 1,6-diphosphate. With respect to the extent of inhibition or activation by the latter compounds, present data suggest the possibility that there are two forms of the cardiac enzyme, one of which is highly sensitive, and a second, which is markedly less sensitive, to the effects of these modifiers.