HUMAN ERYTHROCYTIC CYCLOPHILINS - ISOLATION, CHARACTERIZATION AND STRUCTURE ELUCIDATION

Two cyclosporin A (CsA) binding proteins were purified to homogeneity from human erythrocytes by consecutive steps of repeated gel chromatog raphy, hydrophobic HPLC, ion exchange HPLC and reverse phase HPLC. Pur ified proteins I and II showed almost identical molecular masses and p eptidyl-prolyl cis-trans isomerase (PPIase) activities which were comp letely abolished in the presence of CsA. On the other hand, pi values of the two proteins were distinctly different, 9.1 and 8.1, respective ly, and the CsA-binding affinity (Kd) of protein I was 15.3 nM and tha t of II 25.0 nM. Amino acid sequence analysis of protein I in intact f orm has shown to be Val as the amino terminal residue of the protein, while no phenylthiohydantoinylated amino acid was detected in the sequ ence analysis of intact protein II, suggesting the presence of a modif ied or blocked amino acid at the amino terminus of the protein. The co mplete amino acid sequence of protein I was identified with [Phe(47)]- human cyclophilin A. Significantly high CsA-binding affinities of the present two proteins in human erythrocytes pose a problem of possible implication of CsA bound to such proteins in erythrocytes in pharmacol ogy and/or toxicology of CsA through the blood cell fate.