Spectral magnitude effects on the analyses of secondary structure from circular dichroism spectroscopic data.
The effects of spectral magnitude on the calculated secondary structures derived from circular dichroism (CD) spectra were examined for a number of the most commonly used algorithms and reference databases. Proteins with different secondary structures, ranging from mostly helical to mostly beta-sheet, but which were not components of existing reference databases, were used as test systems. These proteins had known crystal structures, so it was possible to ascertain the effects of magnitude on both the accuracy of determining the secondary structure and the goodness-of-fit of the calculated structures to the experimental data. It was found that most algorithms are highly sensitive to spectral magnitude, and that the goodness-of-fit parameter may be a useful tool in assessing the correct scaling of the data. This means that parameters that affect magnitude, including calibration of the instrument, the spectral cell pathlength, and the protein concentration, must be accurately determined to obtain correct secondary structural analyses of proteins from CD data using empirical methods.
Miles
Andrew J
AJ
Department of Crystallography, Birkbeck College, University of London, London, UK.
Whitmore
Lee
L
Wallace
B A
BA
D000418:Albumins / Q000737:chemistry; D000465:Algorithms; D000818:Animals; D002138:Calibration; D002570:Ceruloplasmin / Q000737:chemistry; D002942:Circular Dichroism / Q000379:methods*; D018360:Crystallography, X-Ray; D019992:Databases as Topic; D030562:Databases, Protein; D024981:Glycogen Phosphorylase / Q000737:chemistry; D006801:Humans; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry*; D011817:Rabbits; D013053:Spectrophotometry / Q000379:methods*; D017356:Synchrotrons
D000418:Albumins; D011506:Proteins; D002570:Ceruloplasmin; D024981:Glycogen Phosphorylase