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A novel anaerobically induced gene of Escherichia coli was identified by insertion of MudI(Amp lac) at approximately 19 minutes on the E. coli chromosome. The gene was tentatively named adhB as it is regulated by adhR, which is the activator gene for adhE. We verified the location of the adhB-lacZ fusion junction via PCR using chromosomal DNA from a strain containing the MudI(Amp lac) insertion. The fusion was found to be in the fourth gene of a putative five gene operon, which is presently being characterized. The first gene of the this operon is ybiK, a putative asparaginase, the second gene is a putative ATP binding protein, and genes three, four (adhB), and five have sequence similarity with the genes dppABC of the E. coli dipeptide permease operon.
Beta-galactosidase assays were performed on strains carrying mutations in regulatory genes as well as the adhB-lacZ fusion. Multiple regulatory genes, including adhR, fnr, modE, cra, crp, csrA, and moe affect expression of adhB. Interestingly, the genes that affect expression of adhB are involved in both fermentation and anaerobic respiration, as well as carbon source regulation. Preliminary sequence data shows that this operon has only a short promoter at the 5'-end. Therefore, we believe that some of the regulators affecting adhB have an indirect effect. To determine which regulators directly affect expression of adhB, we have started by purifying the AdhR protein and will eventually compare the binding of AdhR to the promoter of adhB with that of adhE. The multiple regulatory effects observed suggest that this operon may play an important role in the anaerobic metabolism of Escherichia coli. Additionally, the sequence similarity between the adhB operon and the dpp operon indicate that the adhB operon may encode an amino acid transport system that is important when cells grow anaerobically.
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