Human UDP-Glucuronosyltransferase 1A1 Is the Primary Enzyme Responsible for the N-Glucuronidation of N-Hydroxy-PhIP in Vitro
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- 作者：Michael A. Malfatti and James S. Felton
- 刊名：Chemical Research in Toxicology
- 出版年：2004
- 出版时间：August 2004
- 年：2004
- 卷：17
- 期：8
- 页码：1137 - 1144
- 全文大小：122K
- 年卷期：v.17,no.8(August 2004)
- ISSN：1520-5010
文摘
UDP-glucuronosyltransferase 1A proteins (UGT1A) catalyze the glucuronidation of manyendogenous and xenobiotic compounds including heterocyclic amines and their hydroxylatedmetabolites. Studies have shown that in humans UGT1A-mediated glucuronidation is animportant pathway in the detoxification of food-borne carcinogenic heterocyclic amines. Thebiotransformation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), the most massabundant heterocyclic amine found in cooked meats, is highly dependent on cytochromeP4501A2 hydroxylation followed by UGT-catalyzed glucuronidation of the N-hydroxy-PhIPreactive intermediate. To determine which UGT1A proteins are involved in the glucuronidationof N-hydroxy-PhIP, microsomal preparations from baculovirus-infected insect cells that expressall of the known functional human UGT1A isozymes (UGT1A1, -1A3, -1A4, -1A6, -1A7, -1A8,-1A9, and -1A10) were exposed to N-hydroxy-PhIP and the reaction products were isolated byHPLC. All UGT1A proteins except UGT1A6 showed some degree of activity toward N-hydroxy-PhIP. The formation of both N-hydroxy-PhIP-N2-glucuronide and N-hydroxy-PhIP-N3-glucuronide was both time- and substrate concentration-dependent. UGT1A1 was the most efficientin converting N-hydroxy-PhIP to both conjugates producing five times more of the N2-conjugatethan UGT1A4, the next most active UGT, and 286 times more than UGT1A7, the least activeUGT. With an apparent Km of 52 
r.gif">M and a Kcat of 114 min-1, UGT1A1 was also the mostcatalytically efficient in forming N-hydroxy-PhIP-N2-glucuronide. The catalytic efficiency forN-hydroxy-PhIP-N3-glucuronide formation was 8, 10, and 6 times lower for UGT1A1, -1A4,and -1A8, respectively, when compared to the Kcat values for N-hydroxy-PhIP-N2-glucuronideformation. These results clearly show that UGT1A1 has the highest specificity for glucuronidating N-hydroxy-PhIP. Polymorphic expression resulting in decreased UGT1A1 activityin humans can cause reduced rates of glucuronidation, which can change the metabolic ratiobetween bioactivation and detoxification to favor bioactivation. This change will increase thesusceptibility to the deleterious effects from PhIP exposure because the capacity to form nontoxicN-hydroxy-PhIP glucuronide conjugates will be diminished.
r.gif">M and a Kcat of 114 min-1, UGT1A1 was also the mostcatalytically efficient in forming N-hydroxy-PhIP-N2-glucuronide. The catalytic efficiency forN-hydroxy-PhIP-N3-glucuronide formation was 8, 10, and 6 times lower for UGT1A1, -1A4,and -1A8, respectively, when compared to the Kcat values for N-hydroxy-PhIP-N2-glucuronideformation. These results clearly show that UGT1A1 has the highest specificity for glucuronidating N-hydroxy-PhIP. Polymorphic expression resulting in decreased UGT1A1 activityin humans can cause reduced rates of glucuronidation, which can change the metabolic ratiobetween bioactivation and detoxification to favor bioactivation. This change will increase thesusceptibility to the deleterious effects from PhIP exposure because the capacity to form nontoxicN-hydroxy-PhIP glucuronide conjugates will be diminished.