title: Mechanism of action of lipoprotein lipase on triolein particles: effect of apolipoprotein C-II.
abstract: Triolein particles stabilized by a phosphatidylcholine monolayer were used to study the lipoprotein lipase (LpL) reaction. They were prepared in two different sizes and with triolein and phosphatidylcholine in the molar ratios of 0.9-1.2 : 1 (small particles) and 8-17 : 1 (large particles). The rate of hydrolysis by LpL of phosphatidylcholine on the surface of both lipid particles was only 1/20 as much as that of triolein, even if it was activated to the maximum by apolipoprotein C-II (apoC-II). Thus, the phospholipase activity of LpL was low enough to measure the initial rate of hydrolysis of triolein without causing a gross change of the surface of the lipid particle. When the hydrolysis of triolein by LpL was monitored, fatty acid was released at a constant rate until all of the triolein molecules were hydrolyzed. The enzyme required 220 +/- 17 and 66 +/- 9 nM apoC-II for its half-maximal activity (Km (apoC-II] with small and large particles as a substrate (1.15 mM triolein for small and 2.13 mM triolein for large particles), respectively, using various concentrations of LpL. The Km(apoC-II) values for these two substrates became similar when LpL activity was analyzed with respect to the density of apoC-II on the phosphatidylcholine monolayer at the surface of the particles (bound apoC-II/phosphatidylcholine). The concentration of substrate particles did not affect the Km(apoC-II) values. The presence of an adequate amount of apoC-II increased the maximal activity of LpL (Vmax(triolein)) from 0.48 +/- 0.21 to 6.81 +/- 0.45 and from 0.32 +/- 0.04 to 7.13 +/- 0.64 mmol/h/mg with a slight decrease in the apparent Michaelis constant (Km(triolein)) for small (from 90 to 54 microM triolein) and large (from 1.00 to 0.65 mM triolein) particles, respectively. Although the apparent Km for triolein in large particles was about ten times greater than that in small particles, the values became similar when they were corrected for the concentration of phosphatidylcholine (50-100 microM phosphatidylcholine), which corresponded to the surface area of the substrate particles. It was suggested that bound apoC-II molecules were transferred relatively slowly to other lipid particles while LpL molecules moved rapidly among the lipid particles.(ABSTRACT TRUNCATED AT 400 WORDS)
mesh_terms: D000818:Animals; D053304:Apolipoprotein C-II; D001056:Apolipoproteins C / Q000494:pharmacology*; D002417:Cattle; D006868:Hydrolysis; D007700:Kinetics; D008071:Lipoprotein Lipase / Q000378:metabolism*; D008854:Microscopy, Electron; D010713:Phosphatidylcholines / Q000378:metabolism; D014304:Triolein / Q000378:metabolism*
chemical_list: D053304:Apolipoprotein C-II; D001056:Apolipoproteins C; D010713:Phosphatidylcholines; D014304:Triolein; D008071:Lipoprotein Lipase