Kinetics of Woodward's Reagent K Hydrolysis and Reaction with Staphylococcal Nuclease

The kinetics of the hydrolysis of Woodward's Reagent K has been studied as a function of pH in aqueous solution. Rate constants for the reagent transformation have been combined with rate data for loss of staphylococcal nuclease activity to provide the mechanism for the protein modification. The reactive species for derivatization of the enzyme is the intermediate ketoketenimine. The pH dependence of the rate of the second order reaction between this intermediate and the nuclease yielded a pKapp of 5.22, which is attributed to a carboxyl group or groups of the enzyme.