Peptide Conformations. 46.¹ Conformational Analysis of a Superpotent Cytoprotective Cyclic Somatostatin Analogue

Homo-and heteronuclear 1D-and 2D-NMR techniques have been used to assign proton and carbon resonances of cyclo(-D-Pro-Phe-Thr-Lys(Z)-Trp-Phe-) (1), a potent inhibitor of a transport system in hepatocytes. Independent confirmation of the NMR spectroscopic results was obtained from the analysis of three stereospecifically deuterated isotopomers of 1. The conformational analysis of the backbone is based on evaluation of NOE effects in the rotating frame, chemical shift values, and their temperature dependence, and evaluation of coupling constants. The side-chain conformations are determined from homo-and heteronuclear coupling constants (via DISCO, E.COSY, and COLOC). A βI, βII' structure is deduced for the hydrogen bridges Thr³-CO-HN-Phe⁶ and Phe⁶-CO-HN-Thr³. The derived conformation is refined by two molecular dynamics calculations using the GROMOS program in which the experimental distances obtained from ROE data are included as restraints. Whereas the first calculation (i) uses the conventional force field, the influence of the solvent environment is taken into account in the second calculation (ii) by reducing the charge at the NH protons, which are solvent exposed. The thus calculated conformation (ii) shows minor differences to the original βI, βII' structure. In the conventional MD calculation bifurcated hydrogen bridges with additional γⁱturns are found. In the βI region the Thr hydroxyl group is found to be responsible for the compact packing of 1 because two acceptor hydrogen bonds from the NH protons of Trp⁵ and Phe⁶ and a donor bridge to the carbonyl oxygen of Phe⁶ are formed. The solution structure of 1 was compared with the crystal structure of an analogue of 1 in which Lys(Z) was substituted by a Phe residue. A close similarity of both structures including the orientation of the side chains of the aromatic amino acids was found. However, a distinct difference between the structure in the crystal and in solution is expressed in the side-chain orientation of the Thr residue. The conformation of cyclo(-D-Pro-Phe-Thr-Phe-Trp-Phe-) (7) in Me₂SO solution is also very similar to the conformation of 1.