Abstract

Developing some substitutionary ways such as electrochemical methods might be useful in order to follow structural changes of proteins. In this study, the effect of denaturants such as Urea and SDS (sodium dodecyl sulfate) was investigated on Cytochrome C structure, a metalloproteinase with an oxide/redox property. Structural alterations were studied with potentiometer and different spectrometer techniques. Cytochrome C was prepared in 20mM phosphate buffer with pH=7.0. The protein was treated with different concentrations of Urea and SDS. Related absorbance of native and denatured protein were recorded with Cary 50 (UV-Visible) and AVIV-215 (Circular Dichroism) spectrophotometers in range of 200-700 nm. Electrochemical measurements were also recorded with an acetate/Galvan acetate potentiometer device. Cytochrome C was reported to be structurally stabile in 5-8 and 3-9.5mM concentrations of Urea, which were monitored by voltammetry and spectroscopy methods, respectively. The protein was active in high concentration (26mM) of SDS. The concentration of SDS 20mM seems to be sufficient for proper folding of 0.5mM Cytochrome C. It has been concluded that, 40 mu M Cytochrome C, crudity, and etc do not impact voltammetry results and it is an independent method for detection of structural changes of proteins. Voltammetry method is more sensitive to small changes in protein structure compared with spectroscopy methods.