Effect of proteases on the structure and activity of rabbit skeletal muscle glycogen synthetase

Abstract

Rabbit skeletal muscle glycogen synthetase exists in two forms, the non-phosphorylated, glucose 6-phosphate-independent I-form and the phosphorylated, glucose 6-phosphate-dependent D-form. The conversion of the I-form to the D-form can be effected by phosphorylation catalyzed by either cyclic AMP-dependent [ 1,2] or cyclic AMP-independent [3-51 protein kinases and by limited proteolytic digestion [6-91, whereas conversion brought about by phosphorylation can be reversed by the action of phosphoprotein phosphatase(s), that due to proteases is irreversible. Takeda and Larner [8] and Soderling [9] reported that the conversion of synthetase I to synthetase D by trypsin is accompanied by a decrease in the molecular weight from 90 000 to about 75 000. Takeda and Lamer [8] further reported that the action of trypsin on synthetase is restricted to the COOH-terminal part of the molecule. This report is concerned with further examination of the effect of proteolysis digestion on glycogen synthetase structure and activity using subtilisin as well as trypsin for this purpose. A striking feature of the effect of subtilisin is a marked activation of the enzyme that occurs during the early part of the reaction as seen by activity determinations carried out in the presence or absence of glucose 6-phosphate. In the course of this work the amino acid sequence of the NH2 -terminal portion of the synthetase was determined and found to be slightly different from that reported [lo]. It was noted that low levels of either trypsin or subtilisin cause some modification of the NH2 -terminal region of the synthetase.