title: Alteration of the function of the UDP-glucuronosyltransferase 1A subfamily by cytochrome P450 3A4: different susceptibility for UGT isoforms and UGT1A1/7 variants.
abstract: Functional protein-protein interactions between UDP-glucuronosyltransferase (UGT)1A isoforms and cytochrome P450 (CYP)3A4 were studied. To this end, UGT1A-catalyzed glucuronidation was assayed in Sf-9 cells that simultaneously expressed UGT and CYP3A4. In the kinetics of UGT1A6-catalyzed glucuronidation of serotonin, both Michaelis constant (Km) and maximal velocity (Vmax) were increased by CYP3A4. When CYP3A4 was coexpressed with either UGT1A1 or 1A7, the Vmax for the glucuronidation of the irinotecan metabolite (SN-38) was significantly increased. S50 and Km both which are the substrate concentration giving 0.5 Vmax were little affected by simultaneous expression of CYP3A4. This study also examined the catalytic properties of the allelic variants of UGT1A1 and 1A7 and their effects on the interaction with CYP3A4. Although the UGT1A1-catalyzing activity of 4-methylumbelliferone glucuronidation was reduced in its variant, UGT1A1*6, the coexpression of CYP3A4 restored the impaired function to a level comparable with the wild type. Similarly, simultaneous expression of CYP3A4 increased the Vmax of UGT1A7*1 (wild type) and *2 (N129K and R131K), whereas the same was not observed in UGT1A7*3 (N129K, R131K, and W208R). In the kinetics involving different concentrations of UDP-glucuronic acid (UDP-GlcUA), the Km for UDP-GlcUA was significantly higher for UGT1A7*2 and *3 than *1. The Km of UGT1A7*1 and *3 was increased by CYP3A4, whereas *2 did not exhibit any such change. These results suggest that (1) CYP3A4 changes the catalytic function of the UGT1A subfamily in a UGT isoform-specific manner and (2) nonsynonymous mutations in UGT1A7*3 reduce not only the ability of UGT to use UDP-GlcUA but also CYP3A4-mediated enhancement of catalytic activity, whereas CYP3A4 is able to restore the UGT1A1*6 function.
authors.affiliation: Laboratory of Molecular Life Sciences (Y.Is., H.K., K.K., T.O., Y.Iw., S.T., Y.M., Y.N., N.E., H.Y.) and Laboratory of Medicinal Resource Regulation (F.T., S.M.), Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan; Biotechnology Research Center, Faculty of Engineering, Toyama Prefectural University, Imizu, Toyama, Japan (S.I.); Tohoku Pharmaceutical University, Sendai, Japan (K.N.); Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.Y.); and Department of Clinical Pharmacology, Flinders Medical Centre and Flinders University, Adelaide, Australia (P.I.M.).
authors.forename: Yuji
authors.initials: Y
authors.lastname: Ishii
authors.forename: Hiroki
authors.initials: H
authors.lastname: Koba
authors.forename: Kousuke
authors.initials: K
authors.lastname: Kinoshita
authors.forename: Toshiya
authors.initials: T
authors.lastname: Oizaki
authors.forename: Yuki
authors.initials: Y
authors.lastname: Iwamoto
authors.forename: Shuso
authors.initials: S
authors.lastname: Takeda
authors.forename: Yuu
authors.initials: Y
authors.lastname: Miyauchi
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Nishimura
authors.forename: Natsuki
authors.initials: N
authors.lastname: Egoshi
authors.forename: Futoshi
authors.initials: F
authors.lastname: Taura
authors.forename: Satoshi
authors.initials: S
authors.lastname: Morimoto
authors.forename: Shin'ichi
authors.initials: S
authors.lastname: Ikushiro
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Nagata
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Yamazoe
authors.forename: Peter I
authors.initials: PI
authors.lastname: Mackenzie
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Yamada
mesh_terms: D001711:Biotransformation; D002166:Camptothecin / Q000031:analogs & derivatives / Q000378:metabolism; D002384:Catalysis; D051544:Cytochrome P-450 CYP3A / Q000235:genetics / Q000378:metabolism*; D020719:Glucuronides / Q000378:metabolism*; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006923:Hymecromone / Q000378:metabolism; D007527:Isoenzymes; D007700:Kinetics; D009154:Mutation; D025941:Protein Interaction Mapping; D012701:Serotonin / Q000378:metabolism; D061987:Sf9 Cells; D013379:Substrate Specificity; D014162:Transfection; D000099148:UGT1A1 Enzyme
chemical_list: D002166:Camptothecin; D051544:Cytochrome P-450 CYP3A; D020719:Glucuronides; D014453:Glucuronosyltransferase; D006923:Hymecromone; D007527:Isoenzymes; D012701:Serotonin; D000099148:UGT1A1 Enzyme; C441475:7-ethyl-10-hydroxycamptothecin glucuronide; C510163:CYP3A4 protein, human; C110655:UDP-glucuronosyltransferase, UGT1A6; C524630:UGT1A7 protein, human