Amide modes

Bandekar, J., Amide modes and protein conformation, Biochim Biophys. Acta, 1120, 123, 1992. 

Fig. 5. Comparison of ab initio, DFT/BPW91/6-31G -computed IR and VCD spectra over the amide I, II, and III regions for model peptides (of the generic sequence Ac-Alaw-NHCH3). These are designed to reproduce the major structural features of an o -helix (top left, n— 6, in which the center residue is fully H-bonded), a 3i helix (PLP Il-like, top right, n— 4), and an antiparallel /1-sheet (n= 2, 3 strands, central residue fully H-bonded) in planar (bottom left) and twisted (bottom right) conformations. The computations also encompass all the other vibrations in these molecules, but those from the CH3 side chains were shifted by H/D exchange (CH3) to reduce interference with the amide modes.

Cheam, T. C., and S. Krimm. 1985. Infrared Intensities of Amide Modes in N-methyl-acetamide and Poly(Glycine I) From Ab Initio Calculations of Dipole Moment Derivatives of N-methylacetamide. J. Chem. Phys. 82, 1631-1641. 

In addition, for solid samples or peptides in nonaqueous solvents, the amide II (primarily in-plane NH deformation mixed with C—N stretch, -1500-1530 cm-1) and the amide A (NH stretch, -3300 cm-1 but quite broad) bands are also useful added diagnostics of secondary structure 5,15-17 Due to their relatively broader profiles and complicated by their somewhat weaker intensities, the frequency shifts of these two bands with change in secondary structure are less dramatic than for the amide I yet for oriented samples their polarization properties remain useful 18 Additionally, the amide A and amide II bands are highly sensitive to deuteration effects. Thus, they can be diagnostic of the degree of exchange for a peptide and consequently act as a measure of protected or buried residues as compared to those fully exposed to solvent 9,19,20 Amide A measurements are not useful in aqueous solution due to overlap with very intense water transitions, but amide II measurements can usefully be measured under such conditions 5,19,20 The amide III (opposite-phase NH deformation plus C—N stretch combination) is very weak in the IR and is mixed with other local modes, but has nonetheless been the focus of a few protein-based studies 5,21-26 Finally, other amide modes (IV-VII) have been identified at lower frequencies, but have been the subject of relatively few studies in peptides 5-8,18,27,28 ... 

Vibrational spectroscopy has been used in the past as an indicator of protein structural motifs. Most of the work utilized IR spectroscopy (see, for example, Refs. 118-128), but Raman spectroscopy has also been demonstrated to be extremely useful (129,130). Amide modes are vibrational eigenmodes localized on the peptide backbone, whose frequencies and intensities are related to the structure of the protein. The protein secondary structures must be the main factors determining the force fields and hence the spectra of the amide bands. In particular the amide I band (1600-1700 cm-1), which mainly involves the C=0-stretching motion of the peptide backbone, is ideal for infrared spectroscopy since it has an large transition dipole moment and is spectrally isolated... 

Cheam TC, Krimm S. Infrared intensities of amide modes in N-... 

The concept of transferability and a detailed understanding of these amide modes provides the basis for quantitative estimation of secondary structure for unknown proteins and polypeptides. The quantitative methods currently used to analyze vibrational spectra of proteins can be classified into two categories (1) methods based on decomposition of band contours into underlying components characterized by distinct frequencies, and (2) methods based on principles of pattern recognition. 

Detailed analyses of the vibrational spectra of raacromolecules, however, have provided a deeper understanding of structure and interactions in these systems (Krimm, 1960). An important advance in this direction for proteins came with the determination of the normal modes of vibration of the peptide group in A -methylacetamide (Miyazawa et al., 1958), and the characterization of several specific amide vibrations in polypeptide systems (Miyazawa, 1962, 1967). Extensive use has been made of spectra-structure correlations based on some of these amide modes, including attempts to determine secondary structure composition in proteins (see, for example, Pezolet et al., 1976 Lippert et al., 1976 Williams and Dunker, 1981 Williams, 1983). 

Polypeptide molecules of course exhibit many more vibrational frequencies than the half-dozen or so amide modes. For example, a molecule as simple as the extended form of polyglycine has about 50 bands in its IR and Raman spectra. It is clear that the information contained in the entire spectrum must therefore be a more sensitive indicator of three-dimensional structure. The only way to utilize this information fully is through a normal-mode analysis, that is, by comparing observed frequencies with those calculated for specific secondary structures. This can provide a powerful method for testing structural hypotheses in great detail. 

We illustrate the results of a normal-mode calculation on a small molecule by discussing the normal vibrations of N-methylacetamide (NMA), the simplest molecule containing a trans peptide group analogous to that in a polypeptide chain. A study of this molecule provides insights into the general nature of the so-called amide modes of the peptide group. 

It is worth noting from Table XIII that the approximate force field gives a good reproduction of the frequencies and eigenvectors of the non-CHs modes of the side-chain point-mass model of the a helix. This approximation should therefore be satisfactory for reproducing the amide modes of an a-helical polypeptide chain in, for example, a globular protein. 

A vibrational analysis of a-(GluH) (Sengupta and Krimm, 1985) clearly shows that the side-chain structure does affect the main-chain vibrational modes. The normal modes of a-(GluH) were calculated using the same backbone structure and force field as for a-(Ala) , and a comparison of the results for the amide modes is shown in Table XIV. Some significant differences are observed, and are well predicted, between these two structures. 

Table XIX presents the calculated amide I frequencies and the relative intensities for the A and Ej species modes (expected to be the observed ones) of the and jS - helices. Details for the other amide modes are given in the original paper (Naik and Krimm, 1986a). The nature of the modes differ somewhat between and )3 , being more...

Calculated Amide Mode Frequencies for Various Structures... 

Observed and Calculated Amide Mode Frequencies of CHj-O-Gly—Ala—Ala—Gly—O—CH and Observed Bands of Type I... 

For the IR and Raman spectra, and the detailed description of the normal modes, of this molecule the original publication (Naik and Krimm, 1984a) should be consulted. The 51 Raman and 46 IR bands observed below 1700 cm could be assigned to 68 calculated normal modes with an average error of 6 cm . Comparable assignments could be made for 44 Raman and 50 IR bands observed in this region for the N-deuterated molecule. In Table XXVI we give only the results for the amide modes. 

Observed and Calculated Amide Modes of Type II fi Turns in cyclo(L-Ala—D-Ala-Aca)... 

II. Detailed frequencies and potential energy distributions for both structures are given in the original paper (Bandekar and Krimm, 1985a). In Table XXXII we present the results for the amide modes of the n = 3 molecule we indicate in the discussion the changes when = 5. 

The goal of a vibrational spectroscopic study of a polypeptide molecule is to derive structural information from spectral parameters, such as band frequencies, intensities, and polarizations. In the past, the frequencies of the amide modes were the main diagnostic quantities, with structural insights being obtained from correlational studies based on observed spectra of known polypeptide chain structures. 

We have noted the importance of incorporating calculations of IR intensities in the analysis of spectra. This approach is certain to prove fruitful in a number of areas determination of the dependence of amide mode intensities on conformation influence of size and perfection of structure on intensities correlation of intensities with hydrogen-bond geometry (Cheam and Krimm, 1986). Just as it is possible to develop a conformational (, i/ )-frequency map (Hsu et al., 1976), it should be possible to compute a conformational (, i/ )-intensity map, which could be useful in analyzing the spectra of unordered polypeptide chain structures. Of course, nothing has yet been done on the calculation of Raman intensities of polypeptides, and this area is ripe for future development. 

In the 2D-IR experiments there exists a useful simplification of the description of the spectra when the vibrational dynamics is in the separation of time-scales limit of the so-called Bloch dynamics. Then the correlations of the fluctuations of the various quasi-degenerate amide modes dominate the signals and the interpretations are quite straightforward and analytic. On the basis of experimental determinations of the correlation functions, we explore some of the sensitivities of the 2D-1R signals to the dynamic approximations. In addition, we discuss some of the important possible manipulations of the 2D-IR spectra that permit the display of essentially all possible third-order nonlinear responses from a single data set. 

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