Redox-dependent changes in beta-extended chain and turn structures of cytochrome c in water solution determined by second derivative amide I infrared spectra.
The redox-dependent changes in secondary structure of cytochromes c from horse, cow, and dog hearts in water at 20 degrees C have been determined by amide I infrared spectroscopy. Second derivative amide I spectra were obtained by use of a procedure that includes a convenient method for the effective subtraction of the spectrum of water vapor in the system. The band at 1657 cm-1 representing the helix structure was unaffected by a change in redox state whereas changes in bands due to turns at 1680, 1672, and 1666 cm-1, unordered structure at 1650 cm-1, and beta-structures at 1632 and 1627 cm-1 occurred. About one-fourth of the beta-extended chain spectral region and one-fifth of the beta-turn region (involving a total of approximately 9-13 residues) were sensitive to the oxidation state of heme iron. No significant changes in the secondary structure of either the reduced or oxidized protein due to changes in ionic strength were detected. The localized structural rearrangements triggered by the changes in oxidation state of heme iron are consistent with differences in the binding of heme iron to a histidine imidazole nitrogen and a methionine sulfur atom from the beta-extended chain. The demonstrated ability to obtain highly reproducible second derivative amide I infrared spectra confirms the unique utility of such spectral measurements for localization of subtle changes in secondary structure within a protein, especially for changes among the multiple turns and beta-structures.
Dong
A C
AC
Department of Biochemistry, Colorado State University, Fort Collins 80523.
Huang
P
P
Caughey
W S
WS
D000577:Amides / Q000737:chemistry; D000818:Animals; D002417:Cattle; D003574:Cytochrome c Group / Q000737:chemistry*; D004285:Dogs; D006736:Horses; D009206:Myocardium / Q000201:enzymology; D009994:Osmolar Concentration; D010084:Oxidation-Reduction; D011487:Protein Conformation; D015203:Reproducibility of Results; D012996:Solutions; D013055:Spectrophotometry, Infrared; D013329:Structure-Activity Relationship; D014867:Water
D000577:Amides; D003574:Cytochrome c Group; D012996:Solutions; D014867:Water