Acyl-CoA binding and acylation of UDP-glucuronosyltransferase isoforms of rat liver: their effect on enzyme activity.
When [14C]arachidonoyl-CoA was incubated with crude extracts of rat liver microsomes, [14C]arachidonic acid was incorporated into many proteins, suggesting that modification of these proteins with fatty acid, i.e. acylation, occurred. Using a [14C]arachidonyl-CoA labelling assay, 50 and 53 kDa proteins were purified from rat liver microsomes to near homogeneity by sequential chromatography on Red-Toyopearl, hydroxyapatite, heparin-Toyopearl, Blue-Toyopearl and UDP-hexanolamine-agarose. Acylation of the 50 and 53 kDa proteins occurred in the absence of any other protein, suggesting that these molecules catalyse autoacylation. The acylation was dependent on the length of the incubation period and the concentration of [14C]arachidonoyl-CoA. The 50 and 53 kDa proteins also had acyl-CoA-binding activity; initial rates of acyl-CoA binding and acylation were 0.25 and 0.004 min-1 respectively. The proteins also had weak but distinct acyl-CoA-hydrolysing activity (0.006 min-1). These results suggest that the proteins catalysed the sequential reactions of binding to acyl-CoA, autoacylation, and hydrolysis of fatty acid. N-terminal amino acid sequencing analysis showed these proteins to be UDP-glucuronosyltransferase (UDPGT) isoforms. UDPGT activity was inhibited by arachidonoyl-CoA. These results suggest that binding of acyl-CoA and acylation of UDPGT isoforms regulate the enzyme activities, implying a possible novel function for fatty acyl-CoA in glucuronidation, which is involved in the metabolism of drugs, steroids and bilirubin.
D000214:Acyl Coenzyme A / Q000378:metabolism* / Q000494:pharmacology; D000215:Acylation; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005227:Fatty Acids / Q000737:chemistry; D014453:Glucuronosyltransferase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D008970:Molecular Weight; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013379:Substrate Specificity
D000214:Acyl Coenzyme A; D004791:Enzyme Inhibitors; D005227:Fatty Acids; D007527:Isoenzymes; D011506:Proteins; D016718:Arachidonic Acid; D014453:Glucuronosyltransferase