RT Journal Article
SR Electronic
T1 Influence of N-Terminal Domain Histidine and Proline Residues on the Substrate Selectivities of Human UDP-Glucuronosyltransferase 1A1, 1A6, 1A9, 2B7, and 2B10
JF Drug Metabolism and Disposition
JO Drug Metab Dispos
FD American Society for Pharmacology and Experimental Therapeutics
SP 1948
OP 1955
DO 10.1124/dmd.109.028225
VO 37
IS 9
A1 Oranun Kerdpin
A1 Peter I. Mackenzie
A1 Kushari Bowalgaha
A1 Moshe Finel
A1 John O. Miners
YR 2009
UL http://dmd.aspetjournals.org/content/37/9/1948.abstract
AB An N-terminal domain histidine [corresponding to position 39 of UDP-glucuronosyltransferase (UGT) 1A1] is conserved in all UGT1A and UGT2B subfamily proteins except UGT1A4 (Pro-40) and UGT2B10 (Leu-34). Unlike most UGT1A and UGT2B xenobiotic-metabolizing enzymes, UGT1A4 and UGT2B10 lack the ability to glucuronidate 4-methylumbelliferone (4MU) and 1-naphthol (1NP), both planar phenols, and naproxen (a carboxylic acid). However, only UGT1A4 glucuronidates the tertiary amines lamotrigine (LTG) and trifluoperazine (TFP). In this study, we sought to elucidate the influence of specific N-terminal histidine and proline residues on UGT enzyme substrate selectivity. The conserved N-terminal domain histidine of UGT1A1, UGT1A6, UGT1A9, and UGT2B7 was mutated to proline and leucine 34 of UGT2B10 was substituted with histidine, and the capacity of the wild-type and mutant proteins to glucuronidate 4MU, 1NP, LTG, TFP, and naproxen was characterized. Whereas UGT1A1(H39P), UGT1A6(H38P), and UGT1A9(H37P) lacked the ability to metabolize 4MU, 1NP, and naproxen, all glucuronidated LTG. Km values for UGT1A1(H39P) and UGT1A9(H37P) were 774 and 3812 μM, respectively, compared with 1579 μM for UGT1A4. UGT1A1(H39P) also glucuronidated TFP with a Vmax/Km value comparable to that of UGT1A4. In contrast to the wild-type enzyme, UGT2B10(L34H) glucuronidated 4MU and 1NP with respective Km values of 260 and 118 μM. UGT2B7(H35P) lacked activity toward all substrates. The data confirm a pivotal role for an N-terminal domain proline in the glucuronidation of the tertiary amines LTG and TFP by UGT1A subfamily proteins, whereas glucuronidation reactions involving proton abstraction generally, although not invariably, require a histidine at the equivalent position in both UGT1A and UGT2B enzymes.