ISOLATION AND PURIFICATION OF 2 HUMAN-LIVER UDP-GLUCURONOSYLTRANSFERASES

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Abstract
Two UDP-glucuronosyltransferase (EC 2.4.1.17) were purified from human liver microsomes. Human liver microsomes were solubilized with Emulgen 911 and the UDP-glucuronosyltransferases were separated and purified by chromatofocusing and UDP-hexanolamine Sepharose 4B affinity chromatography. One isoenzyme eluted with an apparent pI of 7.4, displayed a subunit molecular weight of 53,000 after sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and catalyzed the glucuronidation of p-nitrophenol, 4-methylumbelliferone, .alpha.-naphthylamine, and estriol, but not that of 4-aminobiphenyl. A second isoenzyme eluted with an apparent pI of 6.2, displayed a subunit molecular weight of 54,000 after SDS-PAGE, and catalyzed the glucuronidation of p-nitrophenol, 4-methylumbelliferone, .alpha.-naphthylamine, and 4-aminobiphenyl, but not that of estriol. Neither of the purified human liver UDP-glucuronosyltransferases employed estrone, .beta.-estradiol, testosterone, androsterone, or 5.alpha.-androstane-3.alpha., 17.BETA.-diol as substrate. These enzymes displayed apparent Km values in the same order of magnitude for a given substrate. In general, high concentrations of phosphatidylcholine were required for reconstitution of maximal glucuronidation activity. This report documents the existence of multiple UDP-glucuronosyltransferases in human liver.

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