NMR structure of cyclosporin A bound to cyclophilin in aqueous solution

C. Weber, G. Wider, B. Von Freyberg, R. Traber, W. Braun, H. Widmer, K. Wuethrich
1991 Biochemistry  
Cyclosporin A bound to the presumed receptor protein cyclophilin was studied in aqueous solution at p H 6.0 by nuclear magnetic resonance spectroscopy using uniform lSN-or 13C-labeling of cyclosporin A and heteronuclear spectral editing techniques. Sequence-specific assignments were obtained for all but one of the cyclosporin A proton resonances. With an input of 108 intramolecular NOES and four vicinal 3JHNa coupling constants, the three-dimensional structure of cyclosporin A bound to
more » ... A bound to cyclophilin was calculated with the distance geometry program DISMAN, and the structures resulting from 18 1 converged calculations were energy refined with the program FANTOM. A group of 120 conformers was selected on the basis of the residual constraint violations and energy criteria to represent the solution structure. The average of the pairwise root-mean-square distances calculated for the backbone atoms of the 120 structures was 0.58 A. The structure represents a novel conformation of cyclosporin A, for which the backbone conformation is significantly different from the previously reported structures in single crystals and in chloroform solution. The structure has all peptide bonds in the trans form, contains no elements of regular secondary structure and no intramolecular hydrogen bonds, and exposes nearly all polar groups to its environment. The root-mean-square distance between the backbone atoms of the crystal structure of cyclosporin A and the mean of the 120 conformers representing the NMR structure of cyclosporin A bound to cyclophilin is 2.5 I Abbreviations: NMR, nuclear magnetic resonance; CYP, cyclophilin; CsA, cyclosporin A; MeBmt, (4R)-4-[(E)-2-butenyI]-4,N-dimethyl-L-threonine; Abu, L-a-aminobutyric acid; MeLeu, N-methylleucine; MeVal, N-methylvaline; FAB-MS, fast atom bombardment mass spectrometry; hplc, high-performance liquid chromatography; Tris, tris(hydroxymethy1)aminomethane; PMSF, phenylmethanesulfonyl fluoride; EDTA, ethylenediaminetetraacetic acid; DTT, dithiothreitol; MES, 4-morpholineethanesulfonic acid; 1 D, one dimensional; 2D, two dimensional; COSY, two-dimensional correlation spectroscopy; TOCSY, two-dimensional total correlation spectroscopy; NOE, nuclear Overhauser effect; NOESY, two-dimensional NOE spectroscopy; CD, circular dichroism; dAB(iJ) designates the distance between the proton types A and B located in the amino acid residues i and j respectively, where N, NQ, a, and fl denote the amide proton, the N-methyl protons, C"H, and C@I, respectively; RMSD, root-mean-square deviation. Q 1991 American Chemical Societv I ,
doi:10.1021/bi00240a029 pmid:2054355 fatcat:tmbkwwd54rhaflj35voaq3yhbi