Enzymes of the human UDP-glycosyltransferase (UGT) superfamily typically catalyze the covalent addition of the sugar moiety from a UDP-sugar cofactor to relatively low–molecular weight lipophilic compounds. Although UDP-glucuronic acid (UDP-GlcUA) is most commonly employed as the cofactor by UGT1 and UGT2 family enzymes, UGT2B7 and several other enzymes can use both UDP-GlcUA and UDP-glucose (UDP-Glc), leading to the formation of glucuronide and glucoside conjugates. An investigation of UGT2B7-catalyzed morphine glycosidation indicated that glucuronidation is the principal route of metabolism because the binding affinity of UDP-GlcUA is higher than that of UDP-Glc. Currently, it is unclear which residues in the UGT2B7 cofactor binding domain are responsible for the preferential binding of UDP-GlcUA. Here, molecular dynamics (MD) simulations were performed together with site-directed mutagenesis and enzyme kinetic studies to identify residues within the UGT2B7 binding site responsible for the selective cofactor binding. MD simulations demonstrated that Arg259, which is located within the N-terminal domain, specifically interacts with UDP-GlcUA, whereby the side chain of Arg259 H-bonds and forms a salt bridge with the carboxylate group of glucuronic acid. Consistent with the MD simulations, substitution of Arg259 with Leu resulted in the loss of morphine, 4-methylumbelliferone, and zidovudine glucuronidation activity, but morphine glucosidation was preserved.

中文翻译：

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UGT2B7的精氨酸259具有UDP糖选择性

人UDP-糖基转移酶（UGT）超家族的酶通常催化将糖部分从UDP-糖辅因子中共价添加到相对低分子量的亲脂性化合物中。虽然UDP-葡萄糖醛酸（UDP-GlcUA）最常被UGT1和UGT2家族酶用作辅因子，但UGT2B7和其他几种酶可以同时使用UDP-GlcUA和UDP-葡萄糖（UDP-Glc），导致形成葡糖苷酸和葡糖苷结合物。对UGT2B7催化的吗啡糖基化的研究表明，葡糖醛酸化是代谢的主要途径，因为UDP-GlcUA的结合亲和力高于UDP-Glc。目前，尚不清楚UGT2B7辅因子结合结构域中的哪些残基负责UDP-GlcUA的优先结合。这里，分子动力学（MD）模拟与定点诱变和酶动力学研究一起进行，以鉴定UGT2B7结合位点内负责选择性辅因子结合的残基。MD模拟表明，位于N-末端结构域内的Arg259与UDP-GlcUA特异性相互作用，从而Arg259的侧链与葡萄糖醛酸的羧酸酯基团形成氢键并形成盐桥。与MD模拟一致，用Leu取代Arg259导致吗啡，4-甲基伞形酮和齐多夫定葡糖醛酸化活性的丧失，但吗啡糖苷化得以保留。MD模拟表明，位于N-末端结构域内的Arg259与UDP-GlcUA特异性相互作用，从而Arg259的侧链与葡萄糖醛酸的羧酸酯基团形成氢键并形成盐桥。与MD模拟一致，用Leu取代Arg259导致吗啡，4-甲基伞形酮和齐多夫定葡糖醛酸化活性的丧失，但吗啡糖苷化得以保留。MD模拟表明，位于N-末端结构域内的Arg259与UDP-GlcUA特异性相互作用，从而Arg259的侧链与葡萄糖醛酸的羧酸酯基团形成氢键并形成盐桥。与MD模拟一致，用Leu取代Arg259导致吗啡，4-甲基伞形酮和齐多夫定葡糖醛酸化活性的丧失，但吗啡糖苷化得以保留。