Chemical modification of histidyl residues in D-amino acid oxidase from Rhodotorula gracilis.

D-Amino acid oxidase was inactivated by DEP at 30 degrees C and pH 7.5. The reaction followed pseudo-first-order kinetics with a second-order rate constant of 0.254 mM-1.min-1. The pH dependence of the inactivation showed the involvement of a group with a pK of 6.6. The presence of substrate or benzoate protected the enzyme against inactivation. Difference spectra at 242 nm and the reversal of the inactivation in the presence of 1 M hydroxylamine or 0.1 M NaOH pointed to the modification of histidine residues. The statistical analysis of the residual fractional activity versus the number of modified histidine residues led to the conclusion that one histidine residue is essential for the enzyme activity.