title: A synthetic peptide-based assay system for detecting binding between CD36 and an oxidized low-density lipoprotein.
abstract: CD36 is an integral membrane protein that mediates the cellular uptake of oxidized low-density lipoprotein (oxLDL) through recognition of the oxidized glycerophospholipids (oxPLs) formed during LDL oxidation. We aimed to devise an assay system to detect binding between CD36 and oxLDL/oxPL without using recombinant proteins. A peptide corresponding to amino-acid residues 149-168 of mouse CD36 with biotin at its N-terminus (named biotin-CD36(149-168)) and variants of it were synthesized and immobilized onto streptavidin-coated plates. oxLDL labeled with Alexa-Fluor-488 bound specifically and saturably to immobilized biotin-CD36(149-168), but poorly or not at all to the variants, such as that with a scrambled amino-acid sequence. The binding of fluorescence-labeled oxLDL to biotin-CD36(149-168) was inhibited efficiently by an oxPL species, but not by a nonoxidized glycerophospholipid. This assay system using biotin-CD36(149-168) provides a convenient means not only of characterizing binding profiles between CD36 and oxLDL/oxPL but also of finding competitors for the binding.
authors.affiliation: Laboratory of Nutrition Chemistry, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Japan. tkmone@kais.kyoto-u.ac.jp
authors.forename: Satoshi
authors.initials: S
authors.lastname: Tsuzuki
authors.forename: Marie
authors.initials: M
authors.lastname: Takai
authors.forename: Yukari
authors.initials: Y
authors.lastname: Matsuno
authors.forename: Yuki
authors.initials: Y
authors.lastname: Kozai
authors.forename: Maiko
authors.initials: M
authors.lastname: Fujioka
authors.forename: Kozue
authors.initials: K
authors.lastname: Kamei
authors.forename: Hitomi
authors.initials: H
authors.lastname: Inagaki
authors.forename: Ai
authors.initials: A
authors.lastname: Eguchi
authors.forename: Shigenobu
authors.initials: S
authors.lastname: Matsumura
authors.forename: Kazuo
authors.initials: K
authors.lastname: Inoue
authors.forename: Tohru
authors.initials: T
authors.lastname: Fushiki
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D001681:Biological Assay; D001710:Biotin / Q000737:chemistry*; D018955:CD36 Antigens / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry*; D008077:Lipoproteins, LDL / Q000032:analysis* / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000737:chemistry; D011485:Protein Binding
chemical_list: D018955:CD36 Antigens; D005456:Fluorescent Dyes; D008077:Lipoproteins, LDL; D010455:Peptides; C099252:oxidized low density lipoprotein; D001710:Biotin