The 12-residue tryptophan zipper beta-hairpin (SWTWENGKWTWK) and two (13)C-isotopomers were examined in the amide-I region using FTIR and femtosecond two-dimensional infrared (2D IR) spectroscopies. Spectroscopic features of the labeled transitions with (13)C-substituted amide unit present in the terminal or turn region of the hairpin, including their frequency shifts and distributions, line broadenings, orientations, and anharmonicities of diagonal peaks, allow the peptide local structure and local environment to be examined. The results suggest a larger structure fluctuation in the terminal region than in the turn region as a result of the side chain effect and solvent-peptide interaction. The results also suggest that the uncoupled amide-I modes are not degenerate and that this is likely to be a common situation for solvated polypeptides. In addition, the amide-I states in the terminal and turn regions were found to be delocalized over several neighboring amide units. Cross-peaks between the various labeled and unlabeled structural regions were clearly observed in the 2D IR correlation spectra, allowing them to be characterized for monitoring structural changes. These results illustrate the sensitivity of 2D IR to the local environment of solvated peptides. The simulated 1D and 2D IR spectra of the hairpin, obtained by using the vibrational exciton model incorporating coupling constants from quantum chemical computations and semiempirical calculations, were found to reproduce the essential features of the experimental results.

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The principal contributions to the anharmonic coupling of amide vibrations are explored with the objective of comparing recent experiments with density functional theory and evaluating simple models of mode coupling. Experimental information obtained by means of two-dimensional infrared spectroscopy (2D IR) is reasonably well predicted by the computed one- and two-quantum anharmonic modes of amide-A, -I, and -II types in mono-, di- and tripeptides. The expansion of the vibrational energy up to the cubic and quartic coupling of harmonic modes suggested criteria to assess how localized are the forces determining the anharmonicity. The off-diagonal anharmonicity between an amide-A and one other amide mode was shown to be mainly determined by forces involving only these two modes, whereas the off-diagonal anharmonicity of two amide-I modes in peptides depended significantly on forces due to motions other than those of the amide-I type. Both the diagonal and off-diagonal anharmonicities exhibit sensitivity to peptide structures. These results should prove useful in linking 2D IR experimental results to secondary structure. Further, the results are used to evaluate the vibrational exciton model for the mixed-mode anharmonicities of the amide-I transitions.

In this work, we carried out steady-state IR absorption, transient IR pump-probe, and waiting-time-dependent two-dimensional (2D) IR measurements on ferrocyanide and ferricyanide ions solvated in water and deuterated water. These two anions are highly symmetric and have distributed cyano groups with IR-active stretching modes in the 5 μm wavelength region. The line width of their linear IR spectra and the initial value of the vibrational frequency-frequency correlation function extracted from their 2D IR spectra indicate water molecules in the hydration shell of the ferro-species are more inhomogeneously distributed but more tightly bound to the cyano groups than those of the ferri-species. Different charges and their distributions in the two anions cause different hydrogen bonding strengths with solvent. The frequency correlation relaxes somewhat slower in ferrocyanide, agreeing with stronger solute-solvent hydrogen-bonding interaction in this case. Mechanisms of the solvent isotope effect on the vibrational relaxation dynamics of the cyano stretching mode are discussed. These results also suggest that in the hydration shell the ferro-species breaks more water structure than the ferri-species, which is opposite to the situation of the bulk water region (beyond the hydration shell) reported previously. This work demonstrated that combined IR methods can be very useful for understanding the molecular details of the structure and dynamics of the hydrated ions.