Glycogen synthase turnover and phosphorylation in rat H4IIE hepatoma cells

Glycogen synthase was isolated from rat H4IIE hepatoma cells by the use of specific antibodies. Immunoprecipitates from cells grown in the presence of [35S]methionine contained two 35S-labeled polypeptides, designated GS1 and GS2, separable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Labeling of both species was half-maximal after 3 h and remained constant up to 48 h. When cells were incubated with [32P]-phosphate, 32P was incorporated into both species with similar kinetics, half-maximal labeling occurring after 2-3 h. The steady-state ratio 32P/35S was significantly higher for the lower mobility GS2 polypeptide. Pulse-chase experiments showed that the two subunits followed similar kinetics with respect to 35S-labeling. However, the turnover of 32P on the GS2 subunit was significantly faster (t1/2 approximately 30 min) than that on the GS1 subunit (t1/2 approximately 2 h). We suggest that the two polypeptides represent different phosphorylation states of the glycogen synthase subunit and are rapidly interconverted.