Oxidation of pyrrole by dehaloperoxidase-hemoglobin: chemoenzymatic synthesis of pyrrolin-2-ones

Abstract

The use of oxidoreductases as biocatalysts in the syntheses of functionalized, monomeric pyrroles has been a challenge owing to, among a number of factors, undesired polypyrrole formation. Here, we have investigated the ability of dehaloperoxidase (DHP), the coelomic hemoglobin from the terebellid polychaete Amphitrite ornata, to catalyze the H₂O₂-dependent oxidation of pyrroles as a new class of substrate for this enzyme. Substrate oxidation was observed for all compounds employed (pyrrole, N-methylpyrrole, 2-methylpyrrole, 3-methylpyrrole and 2,5-dimethylpyrrole) under both aerobic and anaerobic conditions. Using pyrrole as a representative substrate, only a single oxidation product, 4-pyrrolin-2-one, was observed, and notably without formation of polypyrrole. Reactivity could be initiated from all three biologically relevant oxidation states for this catalytic globin: ferric, ferrous and oxyferrous. Isotope labeling studies determined that the O-atom incorporated into the 4-pyrrolin-2-one product was derived exclusively from H₂O₂, indicative of a peroxygenase mechanism. Consistent with this observation, single- and double-mixing stopped-flow UV-visible spectroscopic studies supported compound I, but not compounds ES or II, as the catalytically-relevant ferryl intermediate involved in pyrrole oxidation. Electrophilic addition of the ferryl oxygen to pyrrole is proposed as the mechanism of O-atom transfer. The results demonstrate the breadth of chemical reactivity afforded by dehaloperoxidase, and provide further evidence for establishing DHP as a multifunctional globin with practical applications as a biocatalyst.