Selective modification of surface-exposed thiol groups in Trigonopsis variabilis D-Amino acid oxidase using poly(ethylene glycol) maleimide and its effect on activity and stability of the enzyme

Covalent modification of purification Trigonopsis variables D-amino acid oxidase using malaimide-activated poly(ethylene glycol) 5000 yielded a stable bioconjugate in which three surface-exposed cysteine side chains were selectively derivatized. Compared with the native enzyme, the PEGylated variant displayed substantially (approximate to 3.3-fold) slowed dissociation rate of FAD cofactor at 50 degrees C, and this caused a twofold thermostabilization of the enzyme activity. The stability under reaction conditions at 30 degrees C was markedly enhance in the PEG-oxidase deamination of D-methionine when 2,6-dichloroindophenol replaced dioxygen as the cosubstrate while it caused a ninefold decrease in substrate catalytic efficiency for the dioxygen-dependant reaction.