K. Oguri et al., PURIFICATION OF A PHENOBARBITAL-INDUCIBLE UDP-GLUCURONOSYLTRANSFERASEISOFORM FROM DOG LIVER WHICH CATALYZES MORPHINE AND TESTOSTERONE GLUCURONIDATION, Archives of biochemistry and biophysics, 325(2), 1996, pp. 159-166
A morphine UDP-glucuronosyltransferase (UGT) which could belong to the
UGT2B subfamily was isolated from liver microsomes of a male beagle d
og treated with phenobarbital, Glucuronidation toward morphine in the
dog liver microsomes was increased threefold by the treatment. The mic
rosomes were solubilized with Emulgen 911 and applied on a column of h
emisuccinate derivative of Sepharose 4B column which has been develope
d in our laboratory, An isoform of UGT in the eluate was purified furt
her by chromatofocusing and UDP-hexanolamine-affinity chromatography,
A purified enzyme, UGT(DOG-PB), was homogeneous on sodium dodecyl sulf
ate polyacrylamide gel electrophoresis and two-dimensional electrophor
esis and exhibited a subunit molecular weight of 50 kDa. This isoform
showed activities toward the 3-hydroxyl group of morphine, 4-hydroxybi
phenyl, 4-nitrophenol, 4-methylumbelliferone, and testosterone, but no
t toward chloramphenicol and the 6-hydroxyl group of morphine. The sub
strate specificity of UGT(DOG-PB) is similar to that of stably express
ed UGT2B1 which is considered a phenobarbital-inducible morphine UGT i
n the rat except that UGT(DOG-PB) is capable of glucuronidating 4-nitr
ophenol but not chloramphenicol. The NH2-terminus until the 30th resid
ue of UGT(DOG-PB) is highly homologous to UGT2B subfamily, and the NH2
-terminal 15 residues of UGT(DOG-PB), are completely identical to thos
e of UGT2B1, UGT2B8, and UGT2B15. This is the first report describing
the UGT isoform of dog and the purification of morphine UGT which may
belong to UGT2B subfamily. (C) 1996 Academic Press, Inc.