An altered-specificity DNA-binding mutant of Escherichia coliσ70 facilitates the analysis
of σ70 function in vivo
Blackwell Science Ltd
Blackwell Publishing Journal Backfiles 1879-2005
The σ subunit of bacterial RNA polymerase is strictly required for promoter recognition. The primary (housekeeping) σ factor of Escherichia coli, σ70, is responsible for most of the gene expression in exponentially growing cells. The fact that σ70 is an essential protein has complicated efforts to genetically dissect the functions of σ70. To facilitate the analysis of σ70 function in vivo, we isolated an altered-specificity DNA-binding mutant of σ70, σ70 R584A, which preferentially recognizes a mutant promoter that is not efficiently recognized by wild-type σ70. Exploiting this σ70 mutant as a genetic tool, we establish an in vivo assay for the inhibitory effect of the bacteriophage T4-encoded anti-σ factor AsiA on σ70-dependent transcription. Our results demonstrate the utility of this altered-specificity system for genetically dissecting σ70 and its interactions with transcription regulators.
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