Assessment of direct interaction between CD36 and an oxidized glycerophospholipid species

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 gamma-hydroxyl(or oxo)-alpha, beta-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.