Purification of Rat-Liver Microsomal UDP-glucuronyltransferase
Purification of Rat-Liver Microsomal UDP-glucuronyltransferase
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大鼠肝微粒体UDP-葡萄糖醛酸转移酶的纯化
DOI：
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发表时间：
1979
期刊：
影响因子：
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通讯作者：
H. Pfeil
中科院分区：
文献类型：
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作者：
K. Bock;D. Josting;W. Lilienblum;H. Pfeil
大鼠肝微粒体UDP-葡萄糖醛酸转移酶催化的葡萄糖醛酸化反应可被3-甲基胆蒽和苯巴比妥诱导。为了阐明这种功能异质性的分子基础，从用诱导剂预处理的大鼠肝脏中纯化酶。
采用胆酸盐增溶，在非离子去污剂Brij 58存在下，在Bio-Gel A-1.5m和DEAE-纤维素上层析，可以分离两种酶形式。随后通过UDP-己醇胺Sepharose 4 B上的亲和层析将两种形式纯化至表观均一。用一种酶形式（酶1）共纯化的3-甲基胆蒽诱导酶对1-萘酚、4-硝基苯酚、3-羟基苯并芘和N-羟基-2-萘胺的活性。相反，苯巴比妥诱导的酶活性对吗啡，氯霉素和4-羟基联苯与其他酶馏分（酶2）。十二烷基硫酸钠/聚丙烯酰胺凝胶显示酶1和酶2的相似的分子量为54000和56000。
结果表明，在大鼠肝脏中存在至少两种形式的UDP-葡萄糖醛酸转移酶。在纯化和膜结合状态的酶活性的影响因素进行了讨论。
Glucuronidation reactions catalysed by rat liver microsomal UDP-glucuronyltransferase are differentially inducible by 3-methylcholanthrene and phenobarbital. To elucidate the molecular basis of this functional heterogeneity the enzyme was purified from livers of rats pretreated with the inducing agents.
Using cholate solubilization, chromatography on Bio-Gel A-1.5m and on DEAE-cellulose in the presence of the nonionic detergent Brij 58, two enzyme forms could be separated. Both forms were subsequently purified to apparent homogeneity by affinity chromatography on UDP-hexanolamine Sepharose 4B. 3-Methylcholanthrene-inducible enzyme activity towards 1-naphthol, 4-nitrophenol, 3-hydroxybenzo(a)pyrene and N-hydroxy-2-naphthylamine copurified with one enzyme form (enzyme 1). In contrast phenobarbital-inducible enzyme activity towards morphine, chloramphenicol and 4-hydroxybiphenyl was associated with the other enzyme fraction (enzyme 2). Sodium dodecylsulfate/polyacrylamide gels showed similar molecular weights of 54000 for enzyme 1 and 56000 for enzyme 2.
The results suggest the presence of at least two forms of UDP-glucuronyltransferase in rat liver. Factors affecting enzyme activity in purified and membrane-bound states are discussed.