RNA polymerase mutants defective in the initiation of transcription-coupled DNA repair

The bacterial Mfd protein is a transcription-repair coupling factor that performs two key functions during transcription-coupled DNA repair. The first is to remove RNA polymerase (RNAP) complexes that have been stalled by a DNA lesion from the site of damage, and the second is to mediate the recruitment of DNA repair proteins. Mfd also displaces transcrip-tion complexes that have been stalled by protein roadblocks, and catalyses the reactivation of tran-scription complexes that have become ‘backtracked’. We have identified amino acid substitutions in the b subunit of Escherichia coli RNAP that disrupt a dir-ect interaction between Mfd and RNAP. These substi-tutions prevent Mfd displacing stalled RNAP from DNA in vivo and in vitro. They define a highly con-served surface-exposed patch on the b1 domain of RNAP that is required by Mfd for the initial step of transcription-coupled repair, the enhancement of roadblock repression and the reactivation of back-tracked transcription complexes