Circular Dichroism Studies on Proteins in Films and in Solution: Estimation of Secondary Structure by g-Factor Analysis

Estimates of the secondary structure of a protein in solution are made by mathematical analyses of its circular dichroism (CD) spectrum below 240 nm. All current procedures require accurate determination of the concentration of the protein sample. Insoluble proteins, such as prions or amyloid, are examined as thin films or gels, but concentrations cannot be precisely defined. The ratio of a sample's CD and absorbance signals is the g-factor, an intensive property. The g-factor spectra of 19 soluble, unconjugated proteins of known structures were measured and used to derive basis spectra, characteristic of the four major structural elements, helix, sheet, turn, and remainder. Using these, the g-factor spectra of other unconjugated proteins, measured in solution or as films, can be analyzed by linear regression to give good estimates of their secondary structures when protein concentration cannot be determined.