The purification and properties of rabbit skeletal muscle glycogen synthase.
The purification and properties of rabbit skeletal muscle glycogen synthase.
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兔骨骼肌糖原合酶的纯化及性质研究
DOI：
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发表时间：
1976
期刊：
影响因子：
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通讯作者：
P. Cohen
中科院分区：
文献类型：
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作者：
H. G. Nimmo;C. Proud;P. Cohen
糖原合成酶a通过内源性磷酸化酶和去支化酶的作用从蛋白质-糖原复合体中溶解，然后经DEAE-纤维素层析，Sepharose4B凝胶过滤或聚乙二醇分级分离，纯化了500倍以上。1000g肌肉在5天内可获得15 mg蛋白质，得率为20%。经凝胶电泳法和超速离心法检测，纯度均在90%以上。测定了氨基酸组成，折射法测得吸光系数A1%280 nm为13.4。糖原合成酶沉淀为两个主要组分，都具有酶活性。较小种(13.3 S)占材料的85%，较大种(19.OS)占15%。经高速沉降平衡离心法测得S组分的相对分子质量为377000。在十二烷基硫酸钠存在下，凝胶电泳法测得亚基相对分子质量为88000，表明13.3-S为四聚体。将该酶的性质与其他工人获得的性质进行了比较。
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 mg 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%280 NM, measured refractiometrically was 13.4. Glycogen synthase a sedimented as two major components, both of which were enzymatically active. The smaller species (13.3 S) comprised 85% and the larger species (19.OS) 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 88 000 indicating that the 13.3-S species is a tetramer. The properties of the enzyme are compared to those obtained by other workers.