ABSTRACT
Lectin-like, oxidized low-density lipoprotein (LDL) receptor 1, LOX-1, is the major receptor for oxidized LDL (OxLDL) in endothelial cells. We have determined the crystal structure of the ligand binding domain of LOX-1, with a short stalk region connecting the domain to the membrane-spanning region, as a homodimer linked by an interchain disulfide bond. In vivo assays with LOX-1 mutants revealed that the "basic spine," consisting of linearly aligned arginine residues spanning over the dimer surface, is responsible for ligand binding. Single amino acid substitution in the dimer interface caused a severe reduction in LOX-1 binding activity, suggesting that the correct dimer arrangement is crucial for binding to OxLDL. Based on the LDL model structure, possible binding modes of LOX-1 to OxLDL are proposed.
Subject(s)
Crystallography, X-Ray , Lipoproteins, LDL/chemistry , Lipoproteins, LDL/metabolism , Receptors, LDL/chemistry , Receptors, LDL/metabolism , Amino Acid Sequence , Amino Acid Substitution , Animals , Arginine/chemistry , Binding Sites , CHO Cells , Conserved Sequence , Cricetinae , Cricetulus , Cysteine/chemistry , Dimerization , Disulfides/chemistry , Humans , Hydrogen Bonding , Ligands , Models, Chemical , Models, Molecular , Molecular Sequence Data , Protein Binding , Protein Folding , Protein Structure, Secondary , Protein Structure, Tertiary , Receptors, LDL/genetics , Receptors, Oxidized LDL , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Scavenger Receptors, Class E , Sequence Homology, Amino AcidABSTRACT
Two different fragments of the ligand-binding domain of LOX-1, the major receptor for oxidized low-density lipoprotein (LDL) on endothelial cells, have been crystallized in different forms. One crystal form contains the disulfide-linked dimer, which is the form of the molecule present on the cell surface; the other contains a monomeric form of the receptor that lacks the cysteine residue necessary to form disulfide-linked homodimers. The crystal of the monomeric ligand-binding domain belongs to space group P2(1)2(1)2(1), with unit-cell parameters a = 56.79, b = 67.57, c = 79.02 A. The crystal of the dimeric form belongs to space group C2, with unit-cell parameters a = 70.86, b = 49.56, c = 76.73 A, beta = 98.59 degrees. Data for the dimeric form of the LOX-1 ligand-binding domain have been collected to 2.4 A. For the monomeric form of the ligand-binding domain, native, heavy-atom derivative and SeMet-derivative crystals have been obtained; their diffraction data have been measured to 3.0, 2.4 and 1.8 A resolution, respectively.
