(442d) Characterization and Comparison of Alkyl Hydroperoxide Reductase and Water-Forming Nadh Oxidase

NADH oxidases are useful biocatalysts for regenerating nicotinamide cofactors of many biological redox reactions. In this presentation, we compare the alkyl hydroperoxide reductase (AhpR) and the H₂O-forming enzyme (nox-2) from Lactococcus lactis (L. lactis), as well as the H₂O-former from Lactobacillus sanfranciscensis (L. sanfranciscensis). AhpR is composed of H₂O₂-forming NADH oxidase (nox-1) and peroxidase. The net reaction of AhpR is the same as nox-2. In this work, both nox-1 and nox-2 are found to be flavoproteins. We found a considerably lower maximum specific activity of nox-1 from L. lactis compared to its nox-2 counterpart or nox-2 from L. sanfranciscensis. Both nox-1 and nox-2 are turnover-limited, as expected for enzymes with labile, redox-active thiols in the active site. In the absence of exogenously added thiols, both nox-1 and nox-1/peroxidase are considerably more stable against overoxidation than nox-2. We will investigate the possibility of using the E. coli-based whole cell system for the oxidative biocatalysis reactions with carbonyl reductase and these flavoproteins.