Purification of three cytosolic glutathione S-transferases from adult Schistosoma mansoni.
Purification of three cytosolic glutathione S-transferases from adult Schistosoma mansoni.
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从成年曼氏血吸虫中纯化三种胞质谷胱甘肽 S-转移酶。
DOI：
10.1016/0003-9861(88)90563-2
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发表时间：
1988
影响因子：
3.9
通讯作者：
Tracy,JW
中科院分区：
文献类型：
--
作者：
O'Leary,KA;Tracy,JW
用原型底物1-氯-2，4-二硝基苯测定，曼氏血吸虫成虫的胞质组分具有谷胱甘肽S-转移酶（EC 2.5.1.18）活性，比在其他后生动物寄生虫中发现的活性高5- 50倍。几个模型底物的调查显示，酶在男性和女性的染色体有不同的，但重叠的底物特异性。四种形式的谷胱甘肽S-转移酶的检测，其中三个，SmGST-1，SmGST-2，和SmGST-3，通过谷胱甘肽亲和层析和HPLC色谱聚焦纯化至表观均一性。纯化的酶表现出非常相似的催化和物理化学性质。它们可以通过对依他尼酸和反式-4-苯基-3-丁烯-2-酮的活性差异来区分，但对芳基卤底物则没有。SmGST-1、SmGST-2和SmGST-3的等电点分别为7.2、7.1和6.9。SmGST-3的多克隆抗血清与其他两种形式的交叉反应，但不与其他可溶性溶酶体蛋白。通过变性条件下的聚丙烯酰胺凝胶电泳，每种纯化的酶显示出28，500的表观亚基分子量。凝胶过滤色谱法得到催化活性形式的酶的分子量为30，800。与所有已知的谷胱甘肽S-转移酶不同，从S. Mansoni似乎是具有催化活性的单体蛋白。
The cytosolic fraction of adultSchistosoma mansonicontains glutathioneS-transferase (EC 2.5.1.18) activity, determined with the prototype substrate 1-chloro-2,4-dinitrobenzene, that is 5- to 50-fold greater than that found in other metazoan parasites. A survey of several model substrates revealed that enzymes in male and female schistosomes have distinct but overlapping substrate specificities. Four forms of glutathioneS-transferase were detected, three of which, SmGST-1, SmGST-2, and SmGST-3, were purified to apparent homogeneity by glutathione affinity chromatography and HPLC chromatofocusing. The purified enzymes displayed very similar catalytic and physicochemical properties. They could be distinguished by differences in activity with ethacrynic acid andtrans-4-phenyl-3-buten-2-one, but not with aryl halide substrates. The isoelectric points of SmGST-1, SmGST-2, and SmGST-3 were estimated to be 7.2, 7.1, 6.9, respectively. A polyclonal antiserum to SmGST-3 cross-reacted with the other two forms, but not with other soluble schistosome proteins. Each of the purified enzymes displayed an apparent subunit molecular weight of 28,500 by polyacrylamide gel electrophoresis under denaturing conditions. Gel filtration chromatography yielded a molecular weight of 30,800 for the catalytically active form of the enzyme. Unlike all known glutathioneS-transferases, the three enzyme forms purified fromS. mansoniappear to be catalytically active monomeric proteins.