Peroxygenase and oxidase activities of dehaloperoxidase‐hemoglobin from Amphitrite ornata (580.1)

Abstract

The marine globin dehaloperoxidase‐hemoglobin (DHP) from Amphitrite ornata was found to catalyze the H2O2‐dependent oxidation of monohaloindoles, a previously unreported class of substrate for DHP. Using 5‐Br‐indole as a representative substrate, the major monooxygenated products were found to be 5‐Br‐2‐oxindole and 5‐Br‐3‐oxindolenine. Isotope labeling studies confirmed that the oxygen atom incorporated was derived exclusively from H2O2, indicative of a previously unreported peroxygenase activity for DHP. Peroxygenase activity could be initiated from either the ferric or oxyferrous states with equivalent substrate conversion and product distribution. It was found that 5‐Br‐3‐oxindole, a precursor of the product 5‐Br‐3‐oxindolenine, readily reduced the ferric enzyme to the oxyferrous state, demonstrating an unusual product‐driven reduction of the enzyme. As such, DHP returns to the globin‐active oxyferrous form after peroxygenase activity ceases. Reactivity with 5‐Br‐3‐oxindole in the absence of H2O2 also yielded 5,5’‐Br2‐indigo above the expected reaction stoichiometry under aerobic conditions, and O2‐concentration studies demonstrated dioxygen consumption. Non‐enzymatic and anaerobic controls both confirmed the requirements for DHP and molecular oxygen in the catalytic generation of 5,5’‐Br2‐indigo, and together suggest a novel oxidase activity for DHP.Grant Funding Source: Supported by the National Science Foundation CHE‐1150709