Microsomal glutathione S-transferase. Purification, initial characterization and demonstration that it is not identical to the cytosolic glutathione S-transferases A, B and C.

R. Morgenstern, C. Guthenberg, J. Depierre
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引用次数: 229

Abstract

Rat liver microsomal glutathione S-transferase was activated with N-ethylmaleimide, solubilized with Triton X-100, and purified by chromatography on hydroxyapatite and CM-Sepharose. A 36-fold purification resulted in a 36% yield, indicating that the glutathione S-transferase accounts for 2.5-3% of the original microsomal protein. The purified protein moved as a band with an apparent molecular weight of 14 000 on sodium dodecyl sulphate gel electrophoresis and appeared to be nearly homogeneous. The complex formed between the purified microsomal glutathione S-transferase and Triton X-100 has a sedimentation coefficient of 3.2 S, a partial specific volume of 0.844 cm3/g, and a Stokes radius of 5.5 nm. The complex has a molecular weight of 127 000 and contains three or four polypeptide chains and 112-134 detergent molecules. Antibodies directed against soluble glutathione S-transferases A, B and C do not react with the purified microsomal enzyme. This finding, together with differences in molecular weight and substrate specificity, demonstrate that the microsomal glutathione S-transferase is an enzyme distinct from the cytosolic glutathione S-transferases.
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微粒体谷胱甘肽s -转移酶。纯化、初步鉴定和证明它与胞质谷胱甘肽s -转移酶A、B和C不相同。
用n -乙基马来酰亚胺活化大鼠肝微粒体谷胱甘肽s -转移酶,用Triton X-100溶解,用羟基磷灰石和CM-Sepharose层析纯化。36倍纯化得到36%的产率,表明谷胱甘肽s转移酶占原始微粒体蛋白的2.5-3%。纯化后的蛋白在十二烷基硫酸钠凝胶电泳上呈带状移动,表观分子量为14000,几乎是均匀的。纯化后的微粒体谷胱甘肽S-转移酶与Triton X-100形成复合物,沉淀系数为3.2 S,部分比容为0.844 cm3/g, Stokes半径为5.5 nm。该配合物的分子量为127000,含有3个或4个多肽链和112-134个洗涤剂分子。针对可溶性谷胱甘肽s -转移酶A、B和C的抗体不与纯化的微粒体酶发生反应。这一发现,再加上分子量和底物特异性的差异,表明微粒体谷胱甘肽s -转移酶是一种不同于胞质谷胱甘肽s -转移酶的酶。
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