The binding of apoA-I-containing ligands to the HDL receptor scavenger receptor class B type I (SR-BI) was characterized using two different assays. The first employed conventional binding or competition assays with (125)I-labeled ligands. The second is a new nonradioactive ligand binding assay, in which the receptor-associated ligand is detected by quantitative immunoblotting ("immunoreceptor assay"). Using both methods, we observed that the K(d) value for spherical HDL (density = 1.1-1.13 g/ml) was approximately 16 microgram of protein/ml, while the values for discoidal reconstituted HDL (rHDL) containing proapoA-I or plasma apoA-I were substantially lower (approximately 0.4-5 microgram of protein/ml). We also observed reduced affinity and/or competition for spherical (125)I-HDL cell association by higher relative to lower density HDL and very poor competition by lipid-free apoA-I and pre-beta-1 HDL. Deletion of either 58 carboxyl-terminal or 59 amino-terminal residues from apoA-I, relative to full-length control apoA-I, resulted in little or no change in the affinity of corresponding rHDL particles. However, rHDL particles containing a double mutant lacking both terminal domains competed poorly with spherical (125)I-HDL for binding to SR-BI. These findings suggest an important role for apoA-I and its conformation/organization within particles in mediating HDL binding to SR-BI and indicate that the NH(2) and COOH termini of apoA-I directly or indirectly contribute independently to binding to SR-BI.
CITATION STYLE
Liadaki, K. N., Liu, T., Xu, S., Ishida, B. Y., Duchateaux, P. N., Krieger, J. P., … Zannis, V. I. (2000). Binding of High Density Lipoprotein (HDL) and Discoidal Reconstituted HDL to the HDL Receptor Scavenger Receptor Class B Type I. Journal of Biological Chemistry, 275(28), 21262–21271. https://doi.org/10.1074/jbc.m002310200
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