Multiple dysfunctions of two apolipoprotein A-I variants, apoA-I (R160L)(Oslo) and apoA-I(P165R), that are associated with hypoalphalipoproteinemia in heterozygous carriers

ApoA-I(R160L)(Oslo) and apoA-I(P165R) are naturally occurring apolipoprotei n (apo) A-I variants that are associated with low HDL-cholesterol in hetero zygous carriers, We characterized the capacity of these variants to bind li pid, to activate lecithin:cholesterol acyltransferase (LCAT), and to promot e efflux of biosynthetic cholesterol from porcine aortic smooth muscle cell s (SMCs) or exogenous cholesterol from lipid-loaded mouse peritoneal macrop hages, During cholate dialysis, normal apoA-I and both variants associated completely with dipalmitoylphosphatidylcholine (DPPC) and formed rLpA-I of identical size. However, both apoA-I(P165R) and apoA-I(R160L)(Oslo) showed a reduced capacity to clear a turbid emulsion of dimyristoylphosphatidylcho line (DMPC), Compared to normal apoA-I, the LCAT-cofactor activity of apoA- I(P165R) and apoA-I(R160L)(Oslo) as defined by the ratio of V-max to appK(m ) was reduced significantly by 62% and 29%, respectively (here and througho ut the text, the apparent K-m is given as Michaelis-Menten kinetics do not take particle binding into account and therefore would result in errors wit h an interfacial enzyme such as LCAT; V-max estimates are not affected by t his error). ApoA-I/DPPC complexes induced biphasic cholesterol efflux from SMCs with a fast and a slow efflux component. Compared to rLpA-I reconstitu ted with wild type apoA-I, rLpA-I with apoA-I(P165R) or apoA-I(R160L)(Oslo) were significantly less effective in promoting cholesterol efflux from SMC s in incubations of 10 min duration but equally effective in incubations of 6 h duration. Lipid-free apoA-I did not induce efflux of biosynthetic chol esterol from SMCs but induced hydrolysis of cholesteryl esters and choleste rol efflux from acetyl-LDG-loaded mouse peritoneal macrophages. In the lipi d-free form, both apoA-I variants promoted normal cholesterol efflux from m urine peritoneal macrophages.jlr We conclude that amino acid residues argin ine 160 and proline 165 of apoA-I contribute to the formation of a domain t hat is very important for initial lipid binding and contributes to LCAT-act ivation and promotion of initial cholesterol efflux but not to the stabiliz ation of preformed rLpA-I.