Protein helical structure determination using CD spectroscopy for solutions with strong background absorbance from 190 to 230nm.

ABSTRACT

Conventional empirical methods for the quantification of the helical content of proteins in solution using circular dichroism (CD) primarily rely on spectral data acquired between wavelengths of 190 and 230nm. The presence of chemical species in a protein solution with strong absorbance within this range can interfere with the ability to use these methods for the determination of the protein's helical structure. The objective of this research was to overcome this problem by developing a method for CD spectral analysis that relies on spectral features above this wavelength range. In this study, we determined that the slopes of CD spectra acquired over the 230 to 240nm region strongly correlate with the helix contents including α-helix and 310-helix of protein as determined using conventional CD algorithms that rely on wavelengths between 190 and 230nm. This approach (i.e., the 230-240nm slope method) is proposed as an effective method to determine the helix content within proteins in the presence of additives such as detergents or denaturants with high absorbance of wavelengths up to 230nm.