The temperature dependence of two kinetic properties of the GT(2P) isoform of microsomal UDP-glucuronosyltransferase was studied for enzyme in intact microsomes and for pure enzyme reconstituted into different types of lipid bilayers. The properties studied were the non-Michaelis-Menten kinetics of the enzyme and activity at V(max(app)). For enzyme in intact microsomes, the pattern of non-Michaelis-Menten kinetics was seen at all temperatures in the range tested (23 to 48°C), and the slopes of the Hill plots of the data were constant across this range of temperatures. Although non-Michaelis-Menten kinetics were present for pure enzyme in bilayers of 1,2-dimyristoylphosphatidylcholine or 1,2-dipalmitoylph osphatidylcholine only in the gel phase (Hockman, Y., Kelley, M., and Zakim, D. (1983) J. Biol. Chem. 258, 6509-6519), it was not possible to reconstitute this pattern of kinetics for enzyme at T > 40°C. For example, GT(2P) displayed Michaelis-Menten kinetics in bilayers of 1,2-distearoylphosphatidylcholine at 44°C. For enzyme in microsomes, activities at V(max(app)) increased with increasing temperature in the range 23 to 48°C, with a discontinuity in the slope of the Arrhenius plot at 34°C. This thermotropic property also could not be reconstituted with pure GT(2P). Instead, activities at V(max(app)) for GT(2P) reconstituted in 1,2-dioleoylphosphatidylcholine, 1,2-distearoylphosphatidylcholine, or 1,2-stearoyl oleoylphosphatidylcholine increased in the range 23 to 37°C, but then decreased at T > 37°C. The fall in activity at T > 37°C was reversible, indicating that GT(2P) undergoes a reversible change at 37°C to a less active form of the enzyme. The differences between pure, reconstituted GT(2P) and microsomal GT(2P) indicate that the thermotropic properties of the microsomal enzyme are not properties of the enzyme per se but depend on interactions between it and other components in the microsome. The data suggest, therefore, that the properties of GT(2P) in microsomes result in part from an organization of components in the microsomal membrane.
|Original language||English (US)|
|Number of pages||5|
|Journal||Journal of Biological Chemistry|
|State||Published - 1989|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology