Assessment of direct interaction between CD36 and an oxidized glycerophospholipid species

Abstract

Cluster of differentiation 36 (CD36) is a transmembrane protein that recognizes multiple diverse ligands. It is believed that (i) oxidized glycerophosphatidylcholine species having a terminal γ-hydroxyl(or oxo)-α,β-unsaturated carbonyl on the sn-2 acyl group (oxGPC_CD36), which can occur on the surface of lipoprotein particles, serve as high-affinity ligands for CD36, and (ii) the amino acid 150–168 of CD36 (CD36_150–168) is responsible for recognizing oxGPC_CD36. However, it remains uncertain whether CD36_150–168 directly interacts with oxGPC_CD36 alone. In this study, we addressed this issue by investigating and comparing the banding pattern by non-denaturing polyacrylamide gel electrophoresis of a glutathione S-transferase (GST) fusion protein containing CD36_150–168 (GST-CD36_150–168), in the presence and absence of an oxGPC_CD36 species, 1-(palmitoyl)-2-(5-keto-6-octenedioyl)phosphatidylcholine (KOdiA-PC). It was shown that GST-CD36_150–168 pre-incubated with KOdiA-PC produced bands at upper positions than did the fusion protein alone. Further analyses revealed that the bands produced by the loading of GST-CD36_150–168/KOdiA-PC mixture represent complexes consisting of the fusion protein and lipid. To our knowledge, this is the first evidence for direct interaction between CD36_150–168 and oxGPC_CD36 alone. It is also notable that the electrophoresis-based technique provides a convenient means to evaluate protein–lipid interactions.

CD36, direct interaction, GST fusion proteins, non-denaturing polyacrylamide gel electrophoresis, oxidized glycerophosphatidylcholine