Involvement of two classes of binding sites in the interactions of cyclophilin B with peripheral blood T-lymphocytes

Cyclophilin B (CyPB) is a cyclosporin A (CsA)-binding protein, mainly assoc iated with the secretory pathway, and is released in biological fluids. We recently reported that CyPB specifically binds to T-lymphocytes and promote s enhanced incorporation of CsA. The interactions with cellular binding sit es involved, at least in part, the specific N-terminal extension of the pro tein. In this study, we intended to specify further the nature of the CyPB- binding sites on peripheral blood T-lymphocytes. We first provide evidence that the CyPB binding to heparin-Sepharose is prevented by soluble sulphate d glycosaminoglycans (GAG), raising the interesting possibility that such i nteractions may occur on the T-cell surface. We then characterized CyPB bin ding to T-cell surface GAG and found that these interactions involved the N -terminal extension of CyPB, but not its conserved CsA-binding domain. In a ddition, we determined the presence of a second CyPB binding site, which we termed a type I site, in contrast with type II for GAG interactions. The t wo binding sites exhibit a similar affinity but the expression of the type I site was 3-fold lower. The conclusion that CyPB binding to the type I sit e is distinct from the interactions with GAG was based on the findings that it was (1) resistant to NaCl wash and GAG-degrading enzyme treatments, (2) reduced in the presence of CsA or cyclophilin C, and (3) unmodified in the presence of either the N-terminal peptide of CyPB or protamine. Finally, w e showed that the type I binding sites were involved in an endocytosis proc ess, supporting the hypothesis that they may correspond to a functional rec eptor for CyPB.