Conformation and calcium-binding properties of a bicyclic nonpeptide

The conformation and calcium binding properties of the bicyclic nonapeptide BCP2, cyclo-(Glu(1)-Ala(2)-Pro(3)-Gly(4)-Lys(5)-Ala(6)-Pro(7)-Gly(8))-cyclo-(1 gamma --> 5 epsilon) Gly(9), have been investigated by means of NMR spectroscopy. Interproton distances, evaluated by, nuclear Overhauser effect (NOE) contacts, and phi angles, from (3)J(NH-alpha CH), have been used to obtain a feasible model for the BCP2-Ca2+ (BCP: bicyclic peptide) complex by means of restrained molecular dynamics (RMD). The NMR analysis of the free peptide, carried out in CD3CN, shows the presence in solution of at least four conformers in intermediate exchange rate. The addition of calcium ions caused the appearance of a new set of resonances, differing from those observed for the free BCP2. A comparison with published data about the conformational behavior of the closely analogous peptide BCP3, differing from BCP2 for two Leu residues instead of two Ala residues in positions 2 and 6, shows that this simple substitution dramatically increases the peptide flexibility. On the contrary, upon calcium ion addition, both BCP2 and BCP3 reach a strictly, close conformation, as strongly testified by the almost identical H-1-NMR spectra exhibited by both peptides. The RMD molecular model of the BCP2-Ca2+ complex, here reported, is a quite symmetric structure, presenting a three-dimensional cavity ideal for the binding of spherical cations. Four carbonyls from the main ring (Ala(2), Gly(4), Ala(6) and Gly(8)) point toward it, offering, together with the two carbonyls of the peptide bridge (Gly(9) and gamma Glu(1)), putative coordinations to the cation