Characterization of the E. Coli Sos Response Protein Ybfe

The transcription of DNA damage response genes in prokaryotes is largely regulated by LexA and controlled through the SOS mechanism. Over 50 genes have been identified as part of the LexA regulon by their upstream LexA binding sequences. Prior work by our lab has shown that loss of the uncharacterized LexA‐regulated gene ybfE is associated with poor survival in E. coli exposed to alkylating agents. In order to elucidate a mechanism, the structure and function of the ybfE gene product were examined. A homology model was built that indicates that YbfE is a DNA‐binding protein that contains a C‐terminal ribbon‐helix‐helix motif and a domain of unknown function at its N‐terminus. The in vivo transcription start site of YbfE is not known, therefore two open reading frames downstream of the SOS operator were examined in vitro for sequence‐specific DNA binding. The first open reading frame contains an additional 23 amino acids at the N‐terminus. Both open reading frames were cloned, expressed, and purified. Electrophoretic mobility shift assays (EMSAs) were performed using the YbfE protein and DNA sequences selected to test binding specificity. Our observations support that YbfE binds DNA nonspecifically in vitro at concentrations above 500 nM. Characterization of sequence specific DNA binding at lower concentrations and protein‐protein interactions is ongoing. Support or Funding Information Supported by American Cancer Society Research Scholar Grant RSG‐12‐161‐01‐DMC.