INFLUENCE OF VESICLE SURFACE-COMPOSITION ON THE INTERFACIAL BINDING OF LECITHIN, CHOLESTEROL ACYLTRANSFERASE AND APOLIPOPROTEIN-A-I

Interfacial binding affinities and capacities of lecithin:cholesterol acyltransferase (LCAT) and apolipoprotein A-I (apoA-I) for surfaces of different phosphatidylcholine (PC) composition, cholesterol content, and apolipoprotein content were measured with a vesicle model system. Native polyacrylamide gel electrophoresis was used to separate Dee pro tein from vesicle-bound protein. ApoA-I was isolated from human plasma and radiolabelled with iodine, whereas radiolabelled LCAT was purifie d from the media of Chinese hamster ovary cells that were transfected with human LCAT cDNA and incubated in the presence of [S-35] cysteine and methionine. Bound and free radiolabelled LCAT and apoA-I and cre q uantified by phosphorimage analysis. ApoA-I binding was not influenced by cholesterol content (14 mole%) but was influenced by the PC fatty acyl composition of the vesicle, PC species containing long chain, pol yunsaturated fatty acids (PUFA) in the sn-2 position resulted in incre ased binding af finity (K-d = 75-177 nM) but reduced capacity (0.1-0.3 apoA-I/1000 PC) in comparison to sn-1 palmitoyl, sn-2 oleoyl PC (POPC , 750 nM and 1.4 apoA-I/1000 PC). LCAT binding affinity to POPC (2190 nM) was stronger in die presence of cholesterol (530 nM), and LCAT bin ding capacity was reduced (2.63 and 0.6 molecules LCAT/1000 PC, respec tively). In comparison to POPC, LCAT binding affinity to sn-1 palmitoy l, sn-2 arachidonyl PC was stronger (611 nM) and binding capacity was reduced (0.7 LCAT/1000 PC). LCAT binding affinity and capacity to sn-1 palmitoyl, sn-2 eicosapentaneoyl PC (2041 nM, and 2.5 LCAT/1000 PC) w ere similar to those observed for POPC. We conclude that vesicle surfa ce PC fatty acyl composition and cholesterol content significantly inf luence LCAT and apoA-I interfacial binding and therefore may alter LCA T enzymatic activity.