LITHIUM-CHLORIDE PERTURBATION OF CIS-TRANS PEPTIDE-BOND EQUILIBRIA - EFFECT ON CONFORMATIONAL EQUILIBRIA IN CYCLOSPORINE-A AND ON TIME-DEPENDENT INHIBITION OF CYCLOPHILIN

The cyclic undecapeptide cyclosporin A (CsA) is a slow-binding inhibitor of the peptidylprolyl cis-trans isomerase (PPIase) cyclophilin. Both the initial inhibitory activity and the subsequent time-dependent inhibition are sensitive to the solvent system (DMSO, THF, LiCl-THF) in which CsA is dissolved prior to the assay. NMR experiments demonstrate that in tetrahydrofuran the MeLeu9-MeLeu10 peptide bond has a cis conformation (Kessler, H.; et al. Biochem. Pharmacol. 1990 40, 169-173). The cis conformer is inactive as a PPIase inhibitor. The same peptide bond adopts a trans conformation when lithium chloride is present as an additive in THF or when CsA is bound to cyclophilin (Fesik, S. W.; et al. Science 1990, 250, 1406-1409). The trans conformer is a tight-binding inhibitor of PPIase activity (K(i) = 20 nM), and the inhibition increases over time (K(i) = 7 nM after 30 min). Detailed kinetic analysis of this transition indicates the presence of at least two interconverting forms of unbound CsA, and a slow structural change in the enzyme-inhibitor complex. The kinetic and structural data taken together suggest that the sequence MeLeu9-trans-MeLeu10-MeVal11 is responsible for efficient binding in the active site of cyclophilin, while the corresponding cis conformer is not recognized at all. The interconversion between the two conformers is kinetically expressed in the time-dependent binding of the drug.