Bradyrhizobium japonicum possesses two discrete sets of electron transfer flavoprotein genes: fixA, fixB and etfS, etfL.

A group of four co-regulated genes (fixA, fixB, fixC, fixX) essential for symbiotic nitrogen fixation has been described in several rhizobial species, including Bradyrhizobium japonicum. The complete nucleotide sequence of the B. japonicum fixA, fixB and fixC, genes is reported here. The derived amino acid sequences confirmed the previously noted sequence similarity between FixA and the beta-subunit and between FixB and the alpha-subunit of mammalian and Paracoccus denitrificans electron transfer flavoproteins (ETF). Since the classical role of ETF is in beta-oxidation of fatty acids, a process unrelated to nitrogen fixation, we rationalized that B. japonicum ought to contain bona fide etf genes in addition to the etf-like genes fixA and fixB. Therefore, we identified, cloned, sequenced, and transcriptionally analyzed the B. japonicum etfSL genes encoding the beta- and alpha-subunits of ETF. The etfSL genes, but not the fix genes, are transcribed in aerobically grown cells. An amino acid sequence comparison between all available ETFs and ETF-like proteins revealed the existence of two distinguishable subfamilies. Group I comprises housekeeping ETFs that link acyl-CoA dehydrogenase reactions with the respiratory chain, such as in the fatty acid degradation pathway. B. japonicum EtfS and EtfL clearly belong to this group. Group II contains ETF-like proteins that are synthesized only under certain specific growth conditions and receive electrons from the oxidation of specific substrates. The products of the anaerobically induced fixA and fixB genes of B. japonicum are members of that group. B. japonicum is the first example of an organism in which genes for proteins of both groups I and II of the ETF family have been identified.