Factors affecting transcriptional regulation of the formate-hydrogen-lyase pathway of Escherichia coli

Abstract

The regulatory elements involved in expression of the gene (fdhF) for the selenopolypeptide of formate dehydrogenase and of a gene (or transcriptional unit) (hyd) specifically responsible for the formation of the gas-evolving hydrogenase (hydrogenase 3) in Escherichia coli were investigated. Formate (or a product of it) is required for expression of both systems since in a pyruvate-formate-lyase deficient mutant induction occurs only when formate is supplemented externally. Under this condition, formate can partially overcome repression by nitrate. The transcription of both the fdhF gene and the hydrogenase-3-encoding systems is independent of the presence of a wild-type fnr gene when formate is present, supporting the view that the Fnr effect on the formation of the formate-hydrogen-lyase pathway is indirect. Mutations blocking the synthesis of a functional molybdenum cofactor also had no major affect on fdhF and hyd expression. The nucleotide sequence of the 5′ flanking region of the fdhF gene was determined and the transcription start point of the fdhF gene was localized by nuclease S1 mapping. Nuclease Bal31 generated deletion clones were constructed and the regulation of their expression was studied. Anaerobic expression and induction by formate depended on the presence of a stretch of approximately 185 nucleotides upstream of the translation start. Elements mediating formate induction and oxygen or nitrate repression could not be separated physically. The regulatory features of the fdhF upstream region bear striking resemblance to systems whose expression are dependent upon upstream activating elements.