A combined spectroscopic and theoretical study of a series of conformationally restricted hexapeptides carrying a rigid binaphthyl-nitroxide donor-acceptor pair

The structural features of a series of linear hexapeptides of general formula Boc-B-Ac-r-A(m)-OtBu, where A is L-Ala or Aib (alpha-aminoisobutyric acid), B is (R)-Bin, a binaphthyl-based C-alpha,C-alpha-disubstituted Gly residue, T is Toac, a nitroxide spin-labeled C-alpha,C-alpha-disubstituted Gly, and r+m = 4, were investigated in methanol solution by fluorescence, transient absorption, IR and CD spectroscopic studies, and by molecular mechanics calculations. These peptides are denoted as B-T/r-m, to emphasize the different position of Toac with respect to that of the Bin fluorophore in the amino acid sequence. The rigidity of the B-T donor - acceptor pair and of the Aib-rich backbone allowed us to investigate the influence of the interchromophoric distance and orientation on the photophysics of the peptides examined. The excited state relaxation processes of binaphthyl were investigated by time-resolved fluorescence and transient absorption experiments. Dynamic quenching of the excited singlet state of binaphthyl by Toac was successfully interpreted by the Forster energy transfer model, provided that the mutual orientation of the chromophores is taken into account. This implies that interconversion among conformational substates, which involves puckering of the Toac piperidine ring, is slow on the time scale of the transfer process, that is slower than 5 ns. By comparison of the experimental and theoretical data, the type of secondary structure (right-handed 3(10) helix) from the B-T/r-m peptides in solution was determined; this would not have been achievable by using the CD and NMR data only, as the data are not diagnostic in this case. Static quenching was observed in all peptides examined but B-T/1-3, where the effect can be ascribed to a non-fluorescent complex. Among the computed low-energy conformers of these peptides, there is one structure exhibiting a NO.-naphthalene center-to-center distance <6 Angstrom, which might be assigned to this complex. The overall results emphasize the versatility of fluorescence experiments in 3D-structural studies in solution.