THE ESCHERICHIA-COLI DSBA GENE IS PARTLY TRANSCRIBED FROM THE PROMOTER OF A WEAKLY EXPRESSED UPSTREAM GENE

The dsbA gene of Escherichia coli encodes a periplasmic enzyme which catalyses disulfide bond formation. Analysis of its surrounding DNA region showed that it is preceded by an open reading frame, orfA, of 984 nucleotides. The intergenic region (19 nucleotides) carries no typical transcription termination signals. dsbA is transcribed from two promoters, the first (91) lies in the distal part of orfA, and the second (P2) just upstream from orfA. Using a plasmid-borne dsbA::TnphoA fusion and an orfA::Omega insertion, each promoter was shown to contribute equally to dsbA transcription, The disruption of the single chromosomal copy of orfA by Omega more drastically reduced the amount of DsbA in the periplasmic space. Such a reduction of the DsbA pool, however, did not change the activities of the AppA, Agp and PhoA periplasmic phosphatases, which all require disulfide bond formation, even when the enzymes were produced from multicopy recombinant plasmids. Thus, in a wildtype strain, DsbA is far from being in limiting amounts for physiological requirements. The orfA gene product was identified as a weakly expressed 39 kDa cytoplasmic protein, but it is not involved in the overall mechanism of disulfide bond formation.