The Purification and Properties of Rabbit Skeletal Muscle Glycogen Synthase

Abstract

Glycogen synthase a was purified over 500‐fold by a procedure which involved solubilisation of the enzyme from a protein‐glycogen complex by the action of endogenous phosphorylase and debranching enzyme, followed by DEAE‐cellulose chromatography, and either gel filtration on Sepharose 4B or fractionation with polyethylene glycol. 15 tng of protein could be obtained from 1000 g of muscle in five days, corresponding to a yield of 20%. The purity was over 90% as judged by gel electrophoresis and ultracentrifugal analysis. The amino acid composition was determined and the absorption coefficient, A1%280nm, measured refractometrically was 13.4. Glycogen synthase a sedimented as two major components, both of which were enzymatically active. The smaller species (13.3S) comprised 85% and the larger species (19.0S) 15% of the material. The molecular weight of the 13.3‐S component was determined to be 377000 by high‐speed sedimentation equilibrium centrifugation. The subunit molecular weight measured by gel electrophoresis in the presence of sodium dodecylsulphate was 88000 indicating that the 13.3‐S species is a tetramer. The properties of the enzyme are compared to those obtained by other workers. Copyright © 1976, Wiley Blackwell. All rights reserved