EFFECT OF THE APOLIPOPROTEIN C-II C-III1 RATIO ON THE CAPACITY OF PURIFIED MILK LIPOPROTEIN-LIPASE TO HYDROLYZE TRIGLYCERIDES IN MONOLAYER VESICLES/

The effect of the apolipoprotein C-II/C-III1 ratio on the capacity of purified bovine milk lipoprotein lipase to hydrolyse triglycerides was measured in a controlled model of pyrene-labeled nonanoyltriglyceride s (1-2 ditetradecyl 3-pyrene nonanoyl glyceride) monolayer vesicles. M onolayer was composed of triglycerides, a non-hydrolysable phospholipi d ether and cholesterol, a model system where the quality of the inter face can be controlled. LPL released fatty acids from pyrene-triglycer ides which were transferred from the lipoprotein structure to albumin. This transfer induces a decrease in the excimer production and in the excimer fluorescence intensity. Apolipoprotein C-II and C-III0, and C -III1 were purified from apolipoprotein VLDL. The 2 fragments, C-III1 A (peptide 1-40) and C-III1 B (peptide 41-79), were obtained after thr ombin cleavage. Apolipoproteins C-III0 and C-IIII1 had a similar inhib itory effect on LPL. Inhibition with apo C-III0 or apo C-III1 was 85% of full LPL activity without inhibitor: Apo C-III1 B inhibited 62% of basal activity. It was 27% less effective than apo C-III1. Fragment C- III1 A did not inhibit LPL. The effect of change in both apo C-II (0-0 .6 mu M) and apo C-III1 (0-1.0 mu M()) on triglyceride hydrolysis show s the importance of. the apo C-II/C-III1 ratio for the release of free fatty acids from triglycerides by LPL. The activating effect of apo C -II in the absence of the apo C-III inhibitor was maximal at 0.06 mu M . No further activation was obtained between 0.06 and 0.30 mu M. Highe r concentrations decreased LPL activity. Apo C-III1 (0.1 mu M) decreas ed the maximum activation by apo C-II from 0.0196 to 0.063 nmol/min/nm ol LPL. Higher concentrations of apo C-III1 (0.1-0.5 mu M) required hi gher apo C-II concentrations (0.30 mu M instead of 0.06 mu M) for maxi mal activation than when apo C-III1 was absent. The activity of the en zyme without apo C-II was decreased by 65% by 0.12 mu M apo C-III1. In creasing the apo C-II/apo C-III1 ratio from 0.1 to 1, increased the ac tivation of the enzyme by a given apo C-II concentration. Moreover, fo r a given apo C-II/C-III1 ratio, the LPL activation increased with the apo C-II concentration (between 0 and 0.010 mu M), until a plateau wa s reached. This is important, as the change in the C-II/C-III1 ratio i s not the only factor affecting LPL activity, and inhibition by apo C- III1 also depends on the overall quantity of apolipoproteins. Extrapol ation of these results suggests that hyperlipoproteinemia seems to be more likely due to overproduction of VLDL, than to a decrease in lipop rotein lipase activity.