Amide I′−II′ 2D IR Spectroscopy Provides Enhanced Protein Secondary Structural Sensitivity

Abstract

We demonstrate how multimode 2D IR spectroscopy of the protein amide I' and II' vibrations can be used to distinguish protein secondary structure. Polarization-dependent amide I'-II' 2D IR experiments on poly-l-lysine in the beta-sheet, alpha-helix, and random coil conformations show that a combination of amide I' and II' diagonal and cross peaks can effectively distinguish between secondary structural content, where amide I' infrared spectroscopy alone cannot. The enhanced sensitivity arises from frequency and amplitude correlations between amide II' and amide I' spectra that reflect the symmetry of secondary structures. 2D IR surfaces are used to parametrize an excitonic model for the amide I'-II' manifold suitable to predict protein amide I'-II' spectra. This model reveals that the dominant vibrational interaction contributing to this sensitivity is a combination of negative amide II'-II' through-bond coupling and amide I'-II' coupling within the peptide unit. The empirically determined amide II'-II' couplings do not significantly vary with secondary structure: -8.5 cm(-1) for the beta sheet, -8.7 cm(-1) for the alpha helix, and -5 cm(-1) for the coil.

Coherent two-dimensional infrared spectroscopy: Quantitative analysis of protein secondary structure in solution

“…More generally, quantifying the spectral signatures of various structural motifs, with particular attention to heterogeneity and site disorder, will further enhance the capabilities of 2DIR spectroscopy as a structural technique. Similarly, the ~12 cm -1 shift that occurs upon deuteration, also suggests that additional information can be derived from the 2DIR of partially deuterated proteins, analogous to amide-I/II 2DIR hydrogen/deuterium exchange experiments, [31] which report directly on the solvent exposure of secondary structures within a protein. Finally, an underlying assumption of the analysis presented here is that the spectra of individual secondary structures are additive.…”

“…The frequency of the delocalized vibrations tends to separate into particular ranges: β-sheets exhibit two peaks near 1630 and 1680 cm -1 whereas α-helices and unstructured regions appear near 1650 cm -1 . [30,31] Despite the secondary structure-sensitivity of the peak frequencies, conformational disorder and solvent exposure render the absorption bands broad and largely featureless.…”

Mapping Molecular Conformations with Multiple-Mode Two-Dimensional Infrared Spectroscopy

“…8À12 2D-IR was developed more or less under such a scenario in the past decade. 13À22 In principle, it is possible to determine the threedimensional molecular conformations by 2D-IR methods: (1) the relative orientations of the chemical bonds of a molecule can be determined by the experimentally determined relative orientations of the transition dipole moments of vibrations; (2) the relative chemical bond distances can be obtained from the experimentally determined vibrational couplings or anharmonicities. Certainly, to realize this ultimate goal, in addition to the advance of experimental techniques, theories and databases to connect the experimental observables and molecular structures are also indispensable.…”

“…Its structures and conformations are displayed in Figure 1. This molecule has several advantages to serve as the model system for the study; (1) it is small enough that the current ab initio calculations can relatively precisely predict many of its molecular properties; (2) it has several possible conformations; (3) it has many typical vibrational groups, for example, CdCÀH, CÀCÀH, CtN, CdC, CdO, and CÀO; (4) these vibrational groups cover all of the molecular space, and therefore, it is possible that the threedimensional conformations of this molecule can be obtained by investigating the vibrations of these groups; and (5) these vibrational groups covers a big vibrational frequency range (>2000 cm À1 ) and a big vibrational spatial separation (more than three chemical bonds), which allows us to explore the sensitivity and potential of our approach.…”

Pump-Probe Spectroscopy, Photon Echoes and Vibrational Wavepackets

“…Two-dimensional IR spectroscopy has been used to examine exciton interactions, internal Stark shifts, and relaxation mechanisms of amide I and II transitions in peptides and proteins [57,59,[82][83][84][85][86][87][88][89][90][91]. Spreading the excitation energies along a second coordinate can separate components that are difficult to resolve in broad one-dimensional IR spectra.…”