diamide and 2-methyl-1-4-hydroquinone

Recently, we showed that the MarR-type repressor YkvE (MhqR) regulates multiple dioxygenases/glyoxalases, oxidoreductases and the azoreductase encoding yvaB (azoR2) gene in response to thiol-specific stress conditions, such as diamide, catechol and 2-methylhydroquinone (MHQ). Here we report on the regulation of the yocJ (azoR1) gene encoding another azoreductase by the novel DUF24/MarR-type repressor, YodB after exposure to thiol-reactive compounds. DNA binding activity of YodB is directly inhibited by thiol-reactive compounds in vitro. Mass spectrometry identified YodB-Cys-S-adducts that are formed upon exposure of YodB to MHQ and catechol in vitro. This confirms that catechol and MHQ are auto-oxidized to toxic ortho- and para-benzoquinones which act like diamide as thiol-reactive electrophiles. Mutational analyses further showed that the conserved Cys6 residue of YodB is required for optimal repression in vivo and in vitro while substitution of all three Cys residues of YodB affects induction of azoR1 transcription. Finally, phenotype analyses revealed that both azoreductases, AzoR1 and AzoR2 confer resistance to catechol, MHQ, 1,4-benzoquinone and diamide. Thus, both azoreductases that are controlled by different regulatory mechanisms have common functions in quinone and azo-compound reduction to protect cells against the thiol reactivity of electrophiles.

Topics: Bacillus subtilis; Bacterial Proteins; Catechols; Cysteine; Diamide; DNA Footprinting; DNA-Binding Proteins; Gene Expression Regulation, Bacterial; Hydrogen Peroxide; Hydroquinones; Mass Spectrometry; Models, Molecular; NADH, NADPH Oxidoreductases; Nitroreductases; Oligonucleotide Array Sequence Analysis; Oxidative Stress; Promoter Regions, Genetic; Proteomics; Quinones; Repressor Proteins; Sulfhydryl Compounds; Transcription, Genetic; Up-Regulation