期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2005
卷号：102
期号：18
页码：6285-6290
DOI：10.1073/pnas.0407872102
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：UDP-glucuronosyltransferase (UGT) isozymes catalyze detoxification of numerous chemical toxins present in our daily diet and environment by conjugation to glucuronic acid. The special properties and enzymatic mechanism(s) that enable endoplasmic reticulum-bound UGT isozymes to convert innumerable structurally diverse lipophiles to excretable glucuronides are unknown. Inhibition of cellular UGT1A7 and UGT1A10 activities and of [33P]orthophosphate incorporation into immunoprecipitable proteins after exposure to curcumin or calphostin-C indicated that the isozymes are phosphorylated. Furthermore, inhibition of UGT phosphorylation and activity by treatment with PKC{epsilon}-specific inhibitor peptide supported PKC involvement. Coimmunoprecipitation, colocalization by means of immunofluorescence, and cross-linking studies of PKC{epsilon} and UGT1A7His revealed that the proteins reside within 11.4 A of each other. Moreover, mutation of three PKC sites in each UGT isozyme demonstrated that T73A/G and T202A/G caused null activity, whereas S432G-UGT1A7 caused a major shift of its pH-8.5 optimum to 6.4 with new substrate selections, including 17{beta}-estradiol. S432G-UGT1A10 exhibited a minor pH shift without substrate alterations. PKC{epsilon} involvement was confirmed by the demonstration that PKC{epsilon} overexpression enhanced activity of UGT1A7 but not of its S432 mutant and the conversion of 17{beta}-[14C]estradiol by S432G-UGT1A7 but not by UGT1A7. Consistent with these observations, treatment of UGT1A7-transfected cells with PKC{epsilon}-specific inhibitor peptide or general PKC inhibitors increased 17{beta}-estradiol catalysis between 5- and 11-fold, with parallel decreases in phosphoserine-432. Here, we report a mechanism involving PKC-mediated phosphorylation of UGT such that phosphoserine/threonine regulates substrate specificity in response to chemical exposures, which possibly confers survival benefit.
