Studies on frog muscle glycogen synthetase. Factors affecting its activity

Abstract

Frog muscle glycogen synthetase is a D form, that is, it depends on glucose 6-P for activity. In this paper data are presented on the effects of Mg2+ and ATP-Mg, as well as the influence of pH on the enzyme activity and stability.

Mg alone produces different effects on crude enzyme extracts or more purified enzyme preparations obtained after 90 minutes of ultracentrifugation at 100.000 X g. In crude extracts it produces an activation time and cation concentration dependent. At high cation concentrations some activity appears without added glucose-6-P, which is parallel to the increase experimented by the P activity. Such Mg concentrations are not physiologycal and this independent-like activity may be due to the small con­ centrations of endogenous glucose-6-P available to the enzyme.

In the purified particulate enzyme Mg2+ always activates strongly without appea­ rance of any independent-like activity. This activation, apparent even without pre­ incubation of the enzyme, can be of 4-5 fold. A Ka of activation of 10-12 mM can be determined in our assay conditions.

Addition of ATP-Mg produces an inactivation of the D-enzyme dependent both on time and concentration of ATP. This inactivation is sensitive to the addition of 3',5'-cycloadenylate at the 10_<s M level. This seems evidence of the action of a kinase that inactivates the D-enzyme by extraphosphorylation.

The optimum pH for enzyme activity seems to be in the range 7.5-7.8 provided the enzyme is left at room temperature or a little higher. At this pH the enzyme has poor stability at temperatures near 0’C, showing maximum activity at pH 6.5 in these conditions. Warming of the enzyme to 30° C at this pH produces an inacti­ vation which is almost total in 60 minutes.