CHARACTERIZATION OF AN ATYPICAL LIPOPROTEIN-BINDING PROTEIN IN HUMAN AORTIC MEDIA MEMBRANES BY LIGAND BLOTTING

By use of ligand-blotting techniques, this study investigated lipoprot ein-binding proteins in human aortic smooth muscle. PAGE was performed under non-reducing conditions, and, using low-density lipoprotein (LD L) as ligand, with rabbit anti-apolipoprotein (ape) B and I-125-labell ed goat anti-rabbit IgG as primary and secondary antibodies respective ly, we demonstrate that membranes from human aortic media (and culture d human smooth-muscle cells) contain a major lipoprotein-binding prote in with an apparent molecular mass of 105 kDa. Anionized preparations (carbamoyl- and acetyl-) of LDL, which did not displace I-125-LDL boun d to the apo B,E receptor of cultured fibroblasts, were also recognize d as ligands for the 105 kDa protein in aortic media membranes. LDL bi nding to 105 kDa protein was decreased in the presence of high density lipoprotein (HDL), although more than 100-fold molar excess of HDL wa s required to achieve 50 % displacement of bound LDL. The LDL-binding activity of 105 kDa protein was inhibited by EDTA, and was also signif icantly decreased when samples were reduced by beta-mercaptoethanol be fore electrophoresis. Monoclonal antibodies against apo B,E receptor r eacted with partially purified bovine adrenal apo B,E receptor, but no t with 105 kDa protein of human aortic media membranes. The spectrum o f properties of this vascular smooth-muscle lipoprotein-binding protei n binding are clearly distinct from those of other previously characte rized lipoprotein-binding molecules.