Circular dichroism protein analysis

J. D. O Neil, K. J. Dorrington, and T. Hofmann, Luminescence and circular-dichroism analysis of terbium binding by pig intestinal calcium-binding protein (relative mass = 9000), Can. J. Biochem. Cell Biol. 62, 434M42 (1984). 

Greenfield, N.J. Circular dichroism analysis for protein-protein interactions. Methods Mol. Biol. 2004, 261, 55-78. 

FT-IR spectroscopy is particularly useful for probing the structure of membrane proteins. Until recently, a lack of adequate experimental techniques has been the reason for the poor understanchng of the secondary structure of most membrane proteins. X-ray diffraction requires high quality crystals and these are not available for many membrane proteins. Circular dichroism (CD) has been widely used for studying the conformation of water-soluble proteins, but problems arise in its use for membrane proteins. The light scattering effect may distort CD spectra and lead to substantial errors in their interpretation. In addition, the reference spectra used for the analysis of CD spectra are based on globular proteins in aqueous solution and may not be applicable to membrane proteins in the hydrophobic environment of lipid bilayers. 

Sreerama, N., Woody, R.W. Computation and analysis of protein circular dichroism spectra. Methods Enzymol. 383, 318-351 (2004)... 

Segel I.H, 1975, Enzyme Kinetics Behmrior and Analysis of Rapid Equilibrium and Steady-State Enzyme Systems, John Wiley Sons, ISBN 0-471-77425-1, Canada Sreerama, N. Woody, R.W. (2004). Computation and analysis of protein circular dichroism spectra. Methods Enzymol, Vol 383, pp 318-351 Stein, R.L., Stiimpler, A.M., Hoii, H. Powers, J.C., (1987). Catalysis by Human Leukocyte Elastase the Proton Inventory as a Mechanistic Probe. Biochemistry, Vol. 26, No.5, pp. 1305-1314... 

Sreerama N, Woody RW 2004. Computation and Analysis of Protein Circular Dichroism Spectra. Methods Enzymology 383 318-351. 

Woody RW, Dunker AK. Aromatic and cysteine sidechain circular dichroism in proteins. In Circular Dichroism and the Conformational Analysis of Biomolecules, ed. GD Fasman. New York Plenum Press, 1996. 

After device construction, structural and functional analysis are critical. One might argue that only the second issue matters, but structural data often give insights into why devices perform suboptimally, and provide important clues about how to improve device function. We routinely use protein analytics (matrix-assisted laser desorption-ionization mass spectroscopy, amino acid composition analysis, gel electrophoresis, Western blotting, circular dichroism, vari-... 

Aside from the direct techniques of X-ray or electron diffraction, the major possible routes to knowledge of three-dimensional protein structure are prediction from the amino acid sequence and analysis of spectroscopic measurements such as circular dichroism, laser Raman spectroscopy, and nuclear magnetic resonance. With the large data base now available of known three-dimensional protein structures, all of these approaches are making considerable progress, and it seems possible that within a few years some combination of noncrystallo-graphic techniques may be capable of correctly determining new protein structures. Because the problem is inherently quite difficult, it will undoubtedly be essential to make the best possible use of all hints available from the known structures. 

Circular dichroism (CD) is a spectroscopic analytical technique used for conformational analysis of peptides and proteins (Johson 1988). It uses the principles of chirality and absorption specifically the different absorption profiles demonstrated by a system for left as opposed to right circularly polarized light. For a system to exhibit CD activity, it must contain a chiral (asymmetric) center that is linked in some way to the chromophore responsible for the absorption. 

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. 

Photoelastic modulation in CD analysis of. see also Circular dichroism proteins, 219, 221-222 Photosynthetic pigment, see Carotenoids Chlorophylls... 

Plane-polarized radiation, CD analysis of proteins, 219-243. see also Circular dichroism Plant cell walls fractionation... 

The coiled-coil motif is an ideal model system for the following reasons there is only one type of secondary structure present (the a-helix) the a-helical structure can be easily monitored by circular dichroism spectroscopy the two-stranded coiled coil contains two subunits stabilized by both intrachain and interchain interactions and, lastly, its small size reduces the potential complexity in the analysis and interpretation of results encountered in the analysis of globular proteins, which have multiple elements of secondary structure (a-helix, (3-sheet, (3-turns, loops, and regions of undefined structure). 

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