Abstract
There is a large discrepancy between the interindividual difference in the hepatic expression level of cytochrome P450 3A4 (CYP3A4) and that of drug clearance mediated by this enzyme. However, the reason for this discrepancy remains largely unknown. Because CYP3A4 interacts with UDP-glucuronosyltransferase 2B7 (UGT2B7) to alter its function, the reverse regulation is expected to modulate CYP3A4-catalyzed activity. To address this issue, we investigated whether protein-protein interaction between CYP3A4 and UGT2B7 modulates CYP3A4 function. For this purpose, we coexpressed CYP3A4, NADPH-cytochrome P450 reductase, and UGT2B7 using a baculovirus-insect cell system. The activity of CYP3A4 was significantly suppressed by coexpressing UGT2B7, and this suppressive effect was lost when UGT2B7 was replaced with calnexin (CNX). These results strongly suggest that UGT2B7 negatively regulates CYP3A4 activity through a protein-protein interaction. To identify the UGT2B7 domain associated with CYP3A4 suppression we generated 12 mutants including chimeras with CNX. Mutations introduced into the UGT2B7 carboxyl-terminal transmembrane helix caused a loss of the suppressive effect on CYP3A4. Thus, this hydrophobic region is necessary for the suppression of CYP3A4 activity. Replacement of the hydrophilic end of UGT2B7 with that of CNX produced a similar suppressive effect as the native enzyme. The data using chimeric protein demonstrated that the internal membrane-anchoring region of UGT2B7 is also needed for the association with CYP3A4. These data suggest that 1) UGT2B7 suppresses CYP3A4 function, and 2) both hydrophobic domains located near the C terminus and within UGT2B7 are needed for interaction with CYP3A4.
Footnotes
- Received February 23, 2015.
- Accepted July 31, 2015.
This study was supported in part by the Japan Research Foundation for Clinical Pharmacology; a Grant-in-Aid for Scientific Research (B) [Grant 25293039] from the Japanese Society of Promotion of Science; and a Grant-in-Aid for Scientific Research (C) [Grant 19590147] from the Ministry of Science, Education, Sports and Technology to Y.I.
This work was presented in part at the 132nd Annual Meeting of the Pharmaceutical Society of Japan, Sapporo, Japan, March 2012 (Miyauchi Y, Ishii Y, Oizaki T, Nagata K, Yamazoe Y, Mackenzie PI, Yamada H, Suppression of cytochrome P450 3A4 function by UDP-glucuronosyltransferase 2B7: Role of C-terminal cytosolic region of UGT2B7); the 19th Microsomes and Drug Oxidation Meeting/12th European Regional Meeting of the International Society for the Study of Xenobiotics, Noordwijk ann Zee, The Netherlands, June 2012 (Miyauchi Y, Ishii Y, Nagata K, Yamazoe Y, Mackenzie PI, Yamada H, UDP-Glucuronosyltransferase (UGT) 2B7 and 1A9 suppress cytochrome P450 3A4 function: evidence for the involvement of the cytosolic tail of UGT in the suppression); the 28th Annual Meeting of the Japanese Society of the Study of Xenobiotics, Tokyo, Japan, October 2013 (Miyauchi Y, Nagata K, Yamazoe Y, Mackenzie PI, Yamada H, Ishii Y, Suppression of cytochrome P450 3A4 activity by UDP-glucuronosyltransferase 2B7: Crucial role of the length of UGT2B7 cytosolic tail in the suppression); the 134th Annual Meeting of the Pharmaceutical Society of Japan, Kumamoto, Japan, March 2014 (Miyauchi Miyauchi Y, Ishii Y, Nagata K, Yamazoe Y, Mackenzie PI, Yamada H, Alteration of cytochrome P450 3A4 activity through a protein-protein interaction with UDP-glucuronosyltransferase 2B7: Cooperation of the luminal domain and cytosolic tail of UGT2B7 is required in the suppression of CYP3A4 function); 19th North American Regional Meeting of the International Society for the Study of Xenobiotics/29th Annual Meeting of the Japanese Society of the Study of Xenobiotics, San Francisco, October, 2014 (Miyauchi Y, Ishii Y, Nagata K, Yamazoe Y, Mackenzie PI, Yamada H, Suppression of cytochrome P450 3A4 activity by UDP-glucuronosyltransferase (UGT) 2B7: the role of charged residue(s) in the cytosolic tail of UGT2B7); and the 20th Microsomes and Drug Oxidation Meeting, Stuttgart, Germany, May, 2014 (Ishii Y, Miyauchi Y, Yamada H, Functional interactions between cytochrome P450 and UDP-glucuronosyltransferase: a new insight into the inter-individual variation of drug metabolism).
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- Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics
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UGT2B7 Suppresses CYP3A4 Function
