PT - JOURNAL ARTICLE AU - Claudia Garrido AU - Silke Leimkuehler TI - The inactivation of human aldehyde oxidase 1 by hydrogen peroxide and superoxide AID - 10.1124/dmd.121.000549 DP - 2021 Jan 01 TA - Drug Metabolism and Disposition PG - DMD-AR-2021-000549 4099 - http://dmd.aspetjournals.org/content/early/2021/06/24/dmd.121.000549.short 4100 - http://dmd.aspetjournals.org/content/early/2021/06/24/dmd.121.000549.full AB - Mammalian aldehyde oxidases (AOX) are molybdoflavo-enzymes of pharmacological and patho-physiological relevance, being involved in phase-I drug metabolism, and also in the generation of reactive oxygen species (ROS) as a product of their enzymatic activity. So far, the physiological role of aldehyde oxidase 1 in the human body remains unknown. The human enzyme hAOX1 is characterized by a broad substrate specificity, oxidizing aromatic/aliphatic aldehydes into their corresponding carboxylic acids and hydroxylating various heteroaromatic rings. The enzyme uses oxygen as terminal electron acceptor to produce hydrogen peroxide and superoxide during turnover. Since hAOX1 and in particular some natural variants not only produce H2O2, but also high amounts of superoxide, we investigated the effect of both ROS molecules on the enzymatic activity of hAOX1 in more detail. We compared hAOX1 to the high O2.- producing natural variant L438V for their time-dependent inactivation with H2O2/O2.- during substrate turnover. We show that the inactivation of the hAOX1 wild type enzyme is mainly based on the production of hydrogen peroxide, while for the variant L438V both hydrogen peroxide and superoxide contribute to the time-dependent inactivation of the enzyme during turnover. Further, the level of inactivation was revealed to be substrate-dependent, using substrates with higher turnover-numbers resulted in a faster inactivation of the enzymes. Analysis of the inactivation site of the enzyme identified a loss of the terminal sulfido-ligand at the molybdenum active site by the produced ROS during turnover. Significance Statement This work characterizes the substrate-dependent inactivation of human aldehyde oxidase 1 under turnover by reactive oxygen species and identifies the site of inactivation. The role of ROS in the inhibition of hAOX1 will have a high impact on future studies.