Mechanism of action of milk lipoprotein lipase at substrate interfaces: effects of apolipoproteins.

The mechanism of action of bovine milk lipoprotein lipase was studied by using a monomolecular film of 1,2-didecanoylglycerol. The apparent rate of hydrolysis of diglyceride increased with increasing surface pressures above 12 mN/m; the enzyme was inactive at pressures less than 12 mN/m. We have measured the effects of four plasma apolipoproteins (apoC-II, apoC-III, apo-I, and apoE), bovine serum albumin, porcine pancreatic colipase, heparin, and NaCl on the kinetics of lipid hydrolysis. At a surface pressure of 15 mN/m, all of the proteins, with the exception of colipase, gave increased enzyme activity compared to lipase alone; apoC-II gave maximal activation. At 25 mN/m, apoC-II at concentrations of less than 0.25 microgram/mL showed a specific activation, whereas the other proteins had no effect. Heparin activated at both high and low surface pressures; NaCl had little or no effect in this system. At a higher concentration of apoC-II (0.50 microgram/mL), the apoprotein inhibited the enzyme. The addition of apoC-III, apoA-I, or apoE (final concentration 0.25 microgram/mL), but not albumin or colipase, to apoC-II (0.25 microgram/mL) caused an increase in surface pressure of 5-6 mN/m and an apparent rate which was less than half that found for lipase alone, suggesting that all of the apoproteins inhibit the apoC-II specific activation.