Using Circular Dichroism Spectra to Estimate Protein Secondary Structure

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.