Characterization of the Mitomycin C Resistance Protein McrA

McrA from Streptomyces lavendulae is a flavin-dependent enzyme thought to provide self-resistance against the DNA-alkylating antibiotic mitomycin C (MMC). McrA belongs to the berberine bridge enzyme (BBE)-like subfamily of oxidases and catalyzes the oxidation of reduced MMC converting it back into the inactive prodrug form, thereby preventing rearrangement into the reactive quinone methide intermediate. Here we demonstrate the first crystal structure of McrA, allowing us to identify key residues involved in retaining MMC in the active site. Biochemical characterization studies such as pre-steady state kinetics verified McrA oxidase activity, while binding studies demonstrated its ability to recognize oxidized MMC.