Identification and expression of the Escherichia coli fdhD and fdhE genes, which are involved in the formation of respiratory formate dehydrogenase

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Abstract
The two closely linked fdhD and fdhE genes of Escherichia coli are required for the formation of active membrane-bound phenazine methosulfate-linked formate dehydrogenase (FDH-PMS). Both genes were isolated from a cosmid library. Restriction endonuclease analysis associated with Mu dII1734 insertion mutagenesis indicated that the two genes were separated by at least 4 kilobases and transcribed in opposite orientations. Initial experiments indicate that the region between the two genes seems not to be essential to FDH-PMS activity. fdhD and fdhE were expressed either in maxicells or from the T7 promoter-polymerase system. They were shown to encode proteins with approximate Mr 30,500 and 32,000, respectively. Both proteins appeared in the soluble fraction and were not recognized by an FDH-PMS-specific antiserum. Therefore, neither fdhD nor fdhE plays a structural role in the formation of FDH-PMS. Expression of a phi(fdhD-lacZ) operon fusion was decreased about threefold by aerobiosis but was indifferent to other effectors tested. It was unaffected by pfl, chlA, selA, and fnr mutations. Expression of a phi(fdhE-lacZ) operon fusion was slightly induced by nitrate. This induction, requiring the presence of functional chl and fnr alleles, was mediated via nitrate metabolism. Transcription of phi(fdhE-lacZ) fusion was fully dependent on wild-type sel alleles. This might suggest the participation of fdhE in the synthesis of the selenopolypeptide of FDH-PMS.