Proteins side chain vibrations

An example of this approach to protein analysis is illustrated by Figure 6.2c, which shows the amide I band of the enzyme lysozyme in D2O. The amide I band of this protein shows nine component bands. The band at 1610 cm is due to amino acid side-chain vibrations and does not contribute to the amide I band. The relative areas of the amide I components are listed in Table 6.2e, and these may be assigned to the various types of secondary structures. The bands at 1623 and 1632 cm... 

Of the modes listed in T able I, one of the most sensitive to conformation in the Raman spectrum is the amide I mode. This band fortunately falls in the range 1630-1700 cm where no other protein bands lie. (The amide III is also very conformationally sensitive, but it is in a frequency region with many side-chain vibrations.) However the liquid water H-O-H bending mode is a broad Raman feature that overlaps with the amide I band so that the contribution of the water must be subtracted out. 

Raman Active Backbone and Side Chain Vibrations of Proteins... 

The vibrational transitions of protein side-chain groups are highly localized (Table 7.7), therefore they can be applied directly to investigate the side chains of peptides and proteins (Singh, 2000). 

The results of these experiments are shown in Figure 9. Of the spectral features apparent in the lgG2a antibody spectram, the most intense band at 1000 cm likely arises from the in-plane ring deformation mode of Phe in IgG (50,51). The amide III protein mode at 1260 cm may be observed in both the IgG and RSV+IgG spectra (52), however, unique, prominent bands are observed in the 1400 - 1600 cm" region, presumably due to selectively enhanced nucleic acid and/or side-chain vibrations (53,54). 

Figure 7.6 Curve-fitted amide I band of myelin basic protein in D2O T, turns and bends R, random coil configuration S, amino acid side-chain vibrations. From Stuart, B., Biological Applications of Infrared Spectroscopy, ACOL Series, Wiley, Chichester, UK, 1997. University of Greenwich, and reproduced by permission of the University of Greenwich.

Side chain vibrational modes are sensitive to the environment and therefore to the conformational variations. Attributions of several bands have been made the S—S and C—S stretching of cystine in the 500-750 cm" range are the most extensively studied. The C—S stretching of Met, the aromatic ring modes of Phe, Tyr, and Trp, the COO" symmetrical stretching, and the CH2 bending modes also have been assigned. These vibrational modes can be used to detect variations of conformation in proteins. 

Near-UV CD of denatured proteins also provides evidence for some order in the side chains, especially in urea- and cold-denatured proteins. Nolting et al (1997) found a broad positive band with possible vibrational... 

In the local mechanical fluctuation model, the local motions of the amino acids on the proximal side of the heme are coupled to the heme through the side group of the proximal histidine. The side chain of the proximal histidine is covalently bonded to the Fe. This bond is the only covalent bond of the heme to the rest of the protein. Thus, motions of the a-helix that contains the proximal histidine are directly coupled the Fe. These motions can push and pull the Fe out of the plane of the heme. Since the CO is bound to the Fe, these motions may induce changes in the CO vibrational transition frequency causing pure dephasing. 

Even where structures of quinone-protein complexes are available from X-ray diffraction experiments, the structures, side-chain conformations, and intermolecular contacts with proteins for the corresponding quinoidal radicals must usually be inferred indirectly from spectroscopic data. The primary spectroscopic methods used to infer structures of quinoidal radicals in photosynthetic reaction center proteins are designed to probe molecular vibrations and spin properties. Directly measurable quantities that are also... 

Proteins, also, can vibrate in part or as a whole in the crystalline state. In contrast to crystals of small molecules, crystals of proteins almost invariably contain large numbers of molecules of solvent of crystallization, often corresponding to 50% or more of the unit cell volume. The extent to which these water molecules are ordered varies dramatically. Near the surface of the protein the water molecules may be well ordered. Beyond the first layer, water molecules typically show increasing levels of disorder. In addition, because of the high solvent content, there may be considerable motion and disorder in the protein molecule, particularly in the orientations of side chains. As a result, the number of independent Bragg reflections that can be measured is reduced, and this effectively reduces the resolution of the electron-density map. 

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