DETERMINATION OF THE NMR SOLUTION STRUCTURE OF THE CYCLOPHILIN-A - CYCLOSPORINE-A COMPLEX

The three-dimensional NMR solution structure of the cyclophilin A (Cyp )-cyclosporin A (CsA) complex was determined, and here we provide a de tailed description of the analysis of the NMR data and the structure c alculation. Using N-15- and C-13-resolved three- and four-dimensional [H-1,H-1]-nuclear Overhauser enhancement (NOE) spectroscopy with unifo rmly isotope-labeled Cyp in the complex, a final data set of 1810 intr a-Cyp, 107 intra-CsA and 63 intermolecular NOE upper distance constrai nts was collected as input for the structure calculation with the prog ram DIANA. A group of DIANA conformers, selected by a previously descr ibed analysis of the dependence of the maximal root-mean-square deviat ion (rmsd) among the individual conformers on the residual target func tion value, was subjected to energy refinement with the program FANTOM . The 22 best energy-refined conformers were then used to represent th e solution structure. The average rmsd relative to the mean structure of these 22 conformers is 1.1 Angstrom for the backbone atoms of all r esidues of the complex. The molecular architecture of Cyp in the Cyp-C sA complex includes an eight-stranded antiparallel beta-barrel, which is closed on each side by an amphipathic helix. CsA is bound in a cavi ty formed by part of the barrel surface and four loops with nonregular secondary structure. Comparison of this structure with structures of Cyp-CsA and other Cyp-peptide complexes determined by different approa ches shows extensive similarities.