RECOMBINANT GST CD36 FUSION PROTEINS DEFINE A THROMBOSPONDIN BINDING DOMAIN - EVIDENCE FOR A SINGLE CALCIUM-DEPENDENT BINDING-SITE ON CD36/

CD36 is a multifunctional cell surface glycoprotein that acts as a sur face receptor for thrombospondin (TSP), and thereby may mediate adhesi ve interactions between cells and substrata, platelets and other cells , and macrophages and apoptotic neutrophils. The identity of the TSP b inding site on CD36 is controversial and may involve more than one str uctural domain. We have constructed a series of recombinant bacterial GST/CD36 fusion proteins that span nearly all of the CD36 molecule and have demonstrated that fusion proteins containing the region extendin g from amino acid 93 to 120 formed specific, saturable, and reversible complexes with TSP. As with intact CD36, binding was calcium-dependen t, was independent of which ligand was immobilized, and was blocked by monoclonal antibodies to both CD36 and TSP. Stoichiometry and affinit y of the fusion proteins for TSP were consistent with that of the inta ct protein. We also demonstrated that these fusion proteins competitiv ely inhibited binding of TSP to puri fied platelet CD36 and to cell su rface CD36 on peripheral blood monocytes and CD36 cDNA-transfected mel anoma cells. These data demonstrate that the region between amino acid s 93 and 120 has all of the characteristics required of the TSP bindin g domain.