Structural studies on rabbit muscle glycogen synthase. II. Limited proteolysis.

Limited tryptic digestion of either synthase I or D forms resulted in the appearance of a new glucose 6-phosphate-dependent form which was composed of 75,000 molecular weight subunits. Early in tryptic digestion, an intermediate 78,000 subunit was also observed with both forms of the enzyme. The NH2-terminal dipeptide sequence of the 75,000 subunit of both forms was the same as that of the original 85,000 subunit (Pro-Leu-) indicating degradation at or near the COOH-terminal end without affecting the NH2-terminal end. Studies interrelating loss of organic phosphate, increase in glucose 6-phosphate dependency, and retention of the NH2-terminal sequence during limited tryptic digestion suggest that there are phosphorylation sites at or near the COOH-terminal, as well as sites within the core of the subunit, which are of importance in the synthase I to D form interconversion reaction via phosphorylation. The pathway of limited tryptic proteolysis of either synthase I or D forms was the same as judged by the molecular weights of the subunit intermediates: 85,000 leads to 78,000 leads to 75,000. A Ca2+-stimulated proteinase activity was also detected in some highly purified preparations of the synthase D form, which led to the appearance of subunits of molecular weight 78,000 and 75,000 together with phosphopeptide(s). These findings suggest that the pathway of proteolysis of the Ca2+-stimulated proteinase is similar to that of trypsin.