Binding of high density lipoprotein (HDL) and discoidal reconstituted HDL to the HDL receptor scavenger receptor class B type I. Effect of lipid association and APOA-I mutations on receptor binding

J Biol Chem. 2000 Jul 14;275(28):21262-71. doi: 10.1074/jbc.M002310200.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Apolipoprotein A-I / chemistry
  • Apolipoprotein A-I / genetics
  • Apolipoprotein A-I / metabolism*
  • Binding Sites
  • CD36 Antigens / chemistry
  • CD36 Antigens / metabolism*
  • Exons
  • Humans
  • Iodine Radioisotopes
  • Kinetics
  • Ligands
  • Lipoproteins, HDL / metabolism*
  • Membrane Proteins*
  • Mice
  • Mutagenesis, Site-Directed
  • Radioligand Assay
  • Receptors, Immunologic*
  • Receptors, Lipoprotein / metabolism
  • Receptors, Scavenger
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Scavenger Receptors, Class B
  • Sequence Deletion
  • Transfection
  • Tumor Cells, Cultured

Substances

  • Apolipoprotein A-I
  • CD36 Antigens
  • Iodine Radioisotopes
  • Ligands
  • Lipoproteins, HDL
  • Membrane Proteins
  • Receptors, Immunologic
  • Receptors, Lipoprotein
  • Receptors, Scavenger
  • Recombinant Proteins
  • SCARB1 protein, human
  • Scarb1 protein, mouse
  • Scavenger Receptors, Class B