Peptides infrared spectroscopy

Hamm P, Urn M and Hochstrasser R M 1998 Structure of the amide I band of peptides measured by femtosecond nonlinear-infrared spectroscopy J. Phys. Chem. B 102 6123-38... 

Gordon, L.M., Lee, K.Y.C., Lipp, M.M., Zasadzinski, J.A., Walther, F.J., Sherman, M. A., and Waring, A.J. Conformational mapping of the N-terminal segment of surfactant protein B in lipid using C-13-enhanced Fourier transform infrared spectroscopy. J. Peptide Res. 

P. Hamm and R. M. Hochstrasser, Structure and dynamics of proteins and peptides Femto second two dimensional infrared spectroscopy, in Ultrafast Infrared and Raman Spectroscopy, Markel Dekker, New York, 2001, p. 273. 

Infrared spectroscopy has been an important part of peptide structural analysis for 50 years now. From a rather basic beginning, applications have blossomed to encompass secondary structure analysis, polarization phenomena, membrane conformation, and orientation, and have extended to time-dependent conformational folding mechanisms. Questions have evolved from basic polymer chemistry to issues centered on peptides involved in socially... 

Molecular weight of heavy and light chains Peptide mapping Amino acid analysis Intrinsic fluorescence spectroscopy Thermal denaturation monitored by fluorescence Fourier transfrome infrared spectroscopy Binding (e.g., ELISA, BiaCore, etc) Potency (e.g., cell based, ELISA)... 

The wavelengths of IR absorption bands are characteristic of specific types of chemical bonds. In the past infrared had little application in protein analysis due to instrumentation and interpretation limitations. The development of Fourier transform infrared spectroscopy (FUR) makes it possible to characterize proteins using IR techniques (Surewicz et al. 1993). Several IR absorption regions are important for protein analysis. The amide I groups in proteins have a vibration absorption frequency of 1630-1670 cm. Secondary structures of proteins such as alpha(a)-helix and beta(P)-sheet have amide absorptions of 1645-1660 cm-1 and 1665-1680 cm, respectively. Random coil has absorptions in the range of 1660-1670 cm These characterization criteria come from studies of model polypeptides with known secondary structures. Thus, FTIR is useful in conformational analysis of peptides and proteins (Arrondo et al. 1993). 

Even though these approaches are powerful methods for determining functional sites on proteins, they are limited if not coupled with some form of structural determination. As Figure 2 illustrates, molecular biology and synthetic peptide/antibody approaches are not only interdependent, they are tied in with structural determination. Structural determination methods can take many forms, from the classic x-ray crystallography and NMR for three-dimensional determination, to two-dimensional methods such as circular dichroism and Fourier Transformed Infrared Spectroscopy, to predictive methods and modeling. A structural analysis is crucial to the interpretation of experimental results obtained from mutational and synthetic peptide/antibody techniques. 

For blood cell membranes the agreement of optical rotatory dispersion and infrared spectroscopy is reassuring. About one-third to one-fourth of the protons in the peptide bonds do not exchange in D20. This non-exchangeable fraction can be equated with the helical content of... 

IR spectroscopy is not a very sensitive analytical tool and is, therefore, not well suited to the detection of small amounts of material. If, however, intermediates have intense and well-resolved IR absorptions, the progress of their chemical transformation can be followed by IR spectroscopy [83,88,91-93], Near-infrared spectroscopy, in combination with an acousto-optic tunable filter, can be sufficiently sensitive to enable the on-bead identification of polystyrene-bound di- and tripeptides, even if the peptides have very similar structures (e.g., Leu-Ala-Gly-PS and Val-Ala-Gly-PS) or differ only in their amino acid sequence (e.g., Leu-Val-Gly-PS and Val-Leu-Gly-PS) [94]. Special resins displaying an IR and Raman barcode have been developed, which may facilitate the deconvolution of combinatorial compound libraries prepared by the mix-and-split method [48]. 

Gonzalez-Martin, I., Hemandez-Hierro, J. M., Vivar-Quintana, A., Revilla, I., and Gonzalez-Perez, C. (2009). The application of near infrared spectroscopy technology and a remote reflectance fibre-optic probe for the determination of peptides in cheeses (cow s, ewe s, and goa t s) with different ripening times. Food Chem. 114,1564—1569. 

Two-Dimensional Coherent Infrared Spectroscopy of Vibrational Excitons in Peptides... 

Structure and Dynamics of Proteins and Peptides Femtosecond Two-Dimensional Infrared Spectroscopy... 

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... 

Asplund MC, Zanni MT, Hochstrasser RM. Two-dimensional infrared spectroscopy of peptides by phase-controlled femtosecond vibrational photon echoes. Proc Natl Acad Sci USA 2000 97 8219-8224. 

Multidimentional nonlinear infrared spectroscopy is used for identification of dynamic structures in liquids and conformational dynamics of molecules, peptides and, in principle, small proteins in solution (Asplund et al., 2000 and references herein). This spectroscopy incorporates the ability to control the responses of particular vibrational transitions depending on their couplings to one another. Two and three-pulse IR photon echo techniques were used to eliminate the inhomogeneous broadening in the IR spectrum. In the third-order IR echo methods, three phase-locked IR pulses with wave vectors kb k2, and k3 are focused on the sample at time intervals. The IR photon echo eventually emitted and the complex 2D IR spectrum is obtained with the use of Fourier transformation. The method was applied to the examination of vibrational properties of N-methyl acetamid and a dipeptide, acyl-proline-NH2.in D20. The 2D IR spectrum showed peaks at 1,610 and 1, 670 cm 1, the two frequencies ofthe acyl-proline dipeptide. Geometry and time-ordering of the incoming pulse sequence in fifth-order 2D spectroscopy is shown in Fig. 1.3. 

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