UDP-glucuronosyltransferase, the role of the amino terminus in dimerization.
UDP-glucuronosyltransferases (UGTs) comprise an important enzyme system in mammals that is involved in detoxification of a variety of small hydrophobic compounds of both endogenous and exogenous origin. Some evidence suggests that these enzymes may function as oligomers; however, little is known about the domain of interaction or the mechanism of oligomerization. In this work, evidence for a functional dimerization between UGTs is provided by studies on mutated forms of UGT2B1. When two inactive forms of UGT2B1 were co-expressed in cell culture, catalytic activity was restored, indicating that UGT2B1 forms functional dimers. To delineate the dimerization domain, inactive fusion proteins containing the amino- or carboxyl-terminal domains of UGT2B1 were generated and expressed with active UGT2B1. Expression of a fusion protein containing only the amino-terminal half of UGT2B1 with active UGT2B1 caused a reduction in UGT2B1 catalytic activity. This reduction in activity was not observed when UGT2B1 was co-expressed with a fusion protein containing only the carboxyl-terminal half of UGT2B1, strongly suggesting that the amino-terminal domain is involved in dimerization. Truncation of the immediate amino terminus of UGT2B1 abolished UGT2B1 activity and dimer formation. Activity was also abolished by an L4R substitution in this region of the mature protein, which is highly conserved in the UGT family. These results indicate that UGTs can interact through their amino-terminal domains to form catalytically active dimers. Possible mechanisms resulting in the formation and stabilization of the UGT2B1 dimer are discussed.
Meech
R
R
Department of Clinical Pharmacology, Flinders University School of Medicine, Bedford Park, South Australia 5042, Australia.
Mackenzie
P I
PI
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002384:Catalysis; D016384:Consensus Sequence; D019281:Dimerization; D014453:Glucuronosyltransferase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D007527:Isoenzymes / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008322:Mammals; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
D007527:Isoenzymes; D046911:Macromolecular Substances; D011993:Recombinant Fusion Proteins; D014453:Glucuronosyltransferase