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Using circular dichroism spectra to estimate protein secondary structure

Nature Protocols volume 1, pages 2876–2890 (2006)Cite this article

Abstract

Circular dichroism (CD) is an excellent tool for rapid determination of the secondary structure and folding properties of proteins that have been obtained using recombinant techniques or purified from tissues. The most widely used applications of protein CD are to determine whether an expressed, purified protein is folded, or if a mutation affects its conformation or stability. In addition, it can be used to study protein interactions. This protocol details the basic steps of obtaining and interpreting CD data, and methods for analyzing spectra to estimate the secondary structural composition of proteins. CD has the advantage that measurements may be made on multiple samples containing ≤20 μg of proteins in physiological buffers in a few hours. However, it does not give the residue-specific information that can be obtained by x-ray crystallography or NMR.

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Figure 1: CD spectra of polypeptides and proteins with representative secondary structures.
Figure 2: Circular dichroism spectra of lysozyme in 10 mM sodium phosphate pH 7.0.

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Acknowledgements

I thank M.A. Andrade, W.C. Johnson, Jr., N. Sreerama, R.W. Woody, S. Venyaminov, and especially the late Gerald D. Fasman for supplying versions of their software for analyzing CD data for use, analysis and distribution. The research was supported by a National Institutes of Health grant, GM-36326 to N.J.G. and to Sarah E. Hitchcock-DeGregori and by the Circular Dichroism Facility at Robert Wood Johnson Medical School (UMDNJ).

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  1. Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, 675 Hoes Lane West, Piscataway, 08854-8021, New Jersey, USA

    Norma J Greenfield

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Greenfield, N. Using circular dichroism spectra to estimate protein secondary structure. Nat Protoc 1, 2876–2890 (2006). https://doi.org/10.1038/nprot.2006.202

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