Chemical Modification of Acidic Residues of Cytochrome c with Safranin: pH Effect on Structure and Function of the Modified Protein

Utilizing cytochrome c (Cyt c), we studied the covalent attachment of safranin O to a protein could affect the protein properties. The effect of covalent attachment of safranin to Cyt c was explored using spectroscopic techniques, which revealed changes in the secondary and tertiary structure of the protein. Using Trp 59 fluorescence emission (lambda(em)=355 nm) and safranin fluorescence profile (lambda(em)=587 nm), we investigated the changes in the structure of the heme moiety and the binding of safranin molecules to the Cyt c protein. Far-UV CD spectroscopy results showed that modification significantly reduced the alpha-helix content. Studies showed that the modification causes the prevention of thermal aggregation of the protein at 65 degrees C and decreases in the peroxidase activity of the protein. Investigating of the pH effect revealed that although the Cyt c modification reduced peroxidase activity, the pH shift to pH 6.0 could increase the peroxidase activity.

Automated summary

By using cytochrome c (Cyt c), the study investigated how the covalent attachment of safranin O affects the properties of a protein. Spectroscopic techniques were employed to explore the impact of covalent attachment on the secondary and tertiary structure of the protein. Results indicated changes in the heme moiety structure and safranin binding to the Cyt c protein through Trp 59 fluorescence emission and safranin fluorescence profile. Far-UV CD spectroscopy revealed a significant reduction in alpha-helix content due to modification. Furthermore, the modification prevented thermal aggregation at 65 degrees C and decreased peroxidase activity, while a shift to pH 6.0 increased the peroxidase activity despite the modification.