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Abstract Bifunctional catalase‐peroxidase (KatG) features a posttranslational methionine‐tyrosine‐tryptophan (MYW) crosslinked cofactor crucial for its catalase function, enabling pathogens to neutralize hydrogen peroxide during infection. We discovered the presence of indole nitrogen‐linked hydroperoxyl adduct (MYW‐OOH) inMycobacterium tuberculosisKatG in the solution state under ambient conditions, suggesting its natural occurrence. By isolating predominantly MYW‐OOH‐containing KatG protein, we investigated the chemical stability and functional impact of MYW‐OOH. We discovered that MYW‐OOH inhibits catalase activity, presenting a unique temporary lock. Exposure to peroxide or increased temperature removes the hydroperoxyl adduct from the protein cofactor, converting MYW‐OOH to MYW and restoring the detoxifying ability of the enzyme against hydrogen peroxide. Thus, theN‐linked hydroperoxyl group is releasable. KatG with MYW‐OOH represents a catalase dormant, but primed, state of the enzyme. These findings provide insight into chemical strategies targeting the bifunctional enzyme KatG in pathogens, highlighting the role ofN‐linked hydroperoxyl modifications in enzymatic function.