Discoidal complexes of A and C apolipoproteins with lipids and their reactions with lecithin: cholesterol acyltransferase.
Micellar, discoidal complexes of human apolipoproteins A-I, A-II, C-I, C-II, C-III-1, and C-III-2 with egg phosphatidylcholine (egg-PC) and cholesterol were prepared by the cholate dialysis method. The complexes, isolated by gel filtration, had similar lipid and protein contents by weight, on the average: 1.77:0.083:1.0, egg-PC/cholesterol/apolipoprotein (w/w). The diameters of the discs, visualized by electron microscopy and estimated by gel filtration, ranged from 100 to 200 A. The alpha-helix content of the apolipoproteins in the complexes was from 50-72%, and their fluorescence properties indicated nonpolar, but quite varied environments for the tryptophan residues in the various complexes. Initial reactions of purified human lecithin: cholesterol acyltransferase with the complexes, adjusted to equal egg-PC concentrations, indicated that all the apolipoproteins activate the enzyme from 6-fold to 400-fold over control vesicles of egg-PC and cholesterol. In decreasing order of reactivity were the complexes with A-I, C-I, C-III-1, C-III-2, C-II, and A-II. These results indicate that aside from lipid-binding capacity and high amphipathic alpha-helix content, other structural features are required for optimal enzyme activation by apolipoproteins. Concentration and temperature dependence experiments gave similar apparent Km values, markedly different apparent Vmax, and very similar activation energies (about 19 kcal/mol), for the various complexes. These observations suggest that the rate-limiting enzymatic step of the reaction is common to all the complexes but that the activated enzyme levels differ from complex to complex. We propose that enzyme activation occurs upon binding to complexes via apolipoproteins. Addition of excess (5-fold) free apolipoprotein A-I or A-II to complexes resulted in the exchange of bound for free apolipoproteins and in loss of reactivity with the enzyme.
D016632:Apolipoprotein A-I; D016633:Apolipoprotein A-II; D001053:Apolipoproteins / Q000378:metabolism*; D001056:Apolipoproteins C; D002784:Cholesterol / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008075:Lipoproteins, HDL / Q000378:metabolism*; D046911:Macromolecular Substances; D008823:Micelles; D007862:Phosphatidylcholine-Sterol O-Acyltransferase / Q000378:metabolism*; D010713:Phosphatidylcholines / Q000378:metabolism*; D013816:Thermodynamics
D016632:Apolipoprotein A-I; D016633:Apolipoprotein A-II; D001053:Apolipoproteins; D001056:Apolipoproteins C; D008075:Lipoproteins, HDL; D046911:Macromolecular Substances; D008823:Micelles; D010713:Phosphatidylcholines; D002784:Cholesterol; D007862:Phosphatidylcholine-Sterol O-Acyltransferase