1887

Abstract

Based on N-terminal sequences obtained from the purified cytoplasmic ferric reductases FerA and FerB, their corresponding genes were identified in the published genome sequence of Pd1222. The and genes were cloned and individually inactivated by insertion of a kanamycin resistance marker, and then returned to for exchange with their wild-type copies. The resulting and mutant strains showed normal growth in brain heart infusion broth. Unlike the mutant, the strain lacking FerA did not grow on succinate minimal medium with ferric 2,3-dihydroxybenzoate as the iron source, and grew only poorly in the presence of ferric sulfate, chloride, citrate, NTA, EDTA and EGTA. Moreover, the mutant strain was unable to produce catechols, which are normally detectable in supernatants from iron-limited wild-type cultures. Complementation of the mutation using a derivative of the conjugative broad-host-range plasmid pEG400 that contained the whole gene and its putative promoter region largely restored the wild-type phenotype. Partial, though significant, restoration could also be achieved with 1 mM chorismate added to the growth medium. The purified FerA protein acted as an NADH : FMN oxidoreductase and catalysed the FMN-mediated reductive release of iron from the ferric complex of parabactin, the major catecholate siderophore of . The deduced amino acid sequence of the FerA protein has closest similarity to flavin reductases that form part of the flavin-dependent two-component monooxygenases. Taken together, our results demonstrate an essential role of reduced flavins in the utilization of exogenous ferric iron. These flavins not only provide the electrons for Fe(III) reduction but most probably also affect the rate of siderophore production.

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2009-04-01
2020-07-09
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