title: Cholesterol-dependent modification of microsomal dynamics and UDPglucuronyltransferase kinetics.
abstract: The effect of both in vitro incorporation and removal of cholesterol in guinea pig liver microsomes on the lipid composition, dynamic properties of the membrane, and kinetic constants of UDPglucuronyltransferase was studied. No significant changes either in the fatty acid composition or in the distribution of phospholipid classes were observed upon cholesterol incorporation and removal. Lateral and rotational mobility measured by the efficiency of pyrene excimer formation and fluorescence of 1,6-diphenylhexatriene decreased with cholesterol incorporation and increased in parallel to cholesterol removal. These changes were associated with alterations in the kinetic properties of UDPglucuronyltransferase. Whereas Vmax increased, the Km of the different steps of the reaction decreased with cholesterol incorporation. The negative homotropic effect and apparent cooperativity of UDP-glucuronic acid decreased when cholesterol was incorporated and increased after cholesterol removal. Moreover, the UDP-N-acetylglucosamine-dependent activation of the enzyme decreased in correlation with an increase of cholesterol concentration in microsomes. It has been demonstrated that both the shift of the non-Michaelian kinetics of the enzyme to Michaelian and the decrease of the UDP-N-acetylglucosamine-dependent activation of the enzyme are evoked by a change of the physical state of the UDPglucuronyltransferase milieu from a gel phase to a liquid-crystalline phase. Therefore, we must admit that cholesterol incorporation in the microsomes while producing an increased packing of the bulk lipids would also cause the separation of more fluid phospholipids, which increase the proportion of molecules in the liquid-crystalline state within the enzyme environment.
mesh_terms: D000818:Animals; D002784:Cholesterol / Q000378:metabolism / Q000494:pharmacology*; D014453:Glucuronosyltransferase / Q000378:metabolism*; D006168:Guinea Pigs; D007700:Kinetics; D008081:Liposomes; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D010743:Phospholipids / Q000378:metabolism
chemical_list: D008081:Liposomes; D010743:Phospholipids; D002784:Cholesterol; D014453:Glucuronosyltransferase