Selective Free Radical Reactions with Proteins and Enzymes: The Inactivation of Ribonuclease

The effect of free radicals derived from thiocyanate, bromide, and carbonate ions on the activity of ribonuclease have been studied in neutral and alkaline solutions. Thiocyanate ions and, to a lesser extent, carbonate ions, protect ribonuclease against radiation-induced inactivation, although the presence of bromide ions has little effect. The absolute rate constants for the reactions of these radical anions with ribonuclease have been measured as a function of pH and salt concentration, and the transient absorption spectra of the reaction products have been determined. The results suggest that damage to a histidine residue, or residues, leads directly to loss of activity and support previous conclusions that exposed tyrosine residues in the enzyme are not essential for activity.