AVIV Model 420 Circular Dichroism Spectrometer


In circular dichroism (CD) spectroscopy, the difference in the absorbances of left- and right-handed circularly polarized light impinging on a solution is measured. Chiral molecules in the solution will absorb one polarization to a greater extent than the other, such that D- and L-tryptophan have CD spectra that are mirror images. For proteins and peptides, the spectra in the far UV region (<260 nm) is usually dominated by the absorbances of the peptide bonds, which has permitted identification of spectra characteristic of various forms of secondary structure found in proteins. This attribute has made CD a standard tool for assessing the conformational integrity of proteins and peptides. Moreover, with its exquisite sensitivity to structural changes, CD is a logical first choice when examining stability and/or the effects of amino acid substitutions. In addition, CD bands above 260 nm originate from the aromatic residues and disulfide bonds, and interactions within their immediate environments. Spectra in this region can provide a fingerprint of the correctly folded conformation of a protein that is surpassed in sensitivity only by NMR spectroscopy. When characterizing the effects of mutations, CD is an ideal tool for ascertaining conformational stability and/or the kinetics of changes in response to temperature or upon additions to the solvent.

In the Aviv instrument, CD and absorbance spectra can be recorded simultaneously from 170 to 875 nm with temperature control for up to five samples from -5 to 110 °C. Other accessories permit measurement of total fluorescence along with CD and automated titrations. The programmability of the instrument and its accessories make sophisticated multidimensional experiments feasible.For a brief overview of spectroscopic tools for structural studies including CD, FTIR, Raman, and NMR, see: Pelton, J. T. and McLean, L. R. (2000) Spectroscopic methods for analysis of protein secondary structure. Anal. Biochem. 277, 167-176.

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