Distribution of nitrogen fixation and nitrogenase-like sequences amongst microbial genomes

Abstract

BackgroundThe metabolic capacity for nitrogen fixation is known to be present in several prokaryotic species scattered across taxonomic groups. Experimental detection of nitrogen fixation in microbes requires species-specific conditions, making it difficult to obtain a comprehensive census of this trait. The recent and rapid increase in the availability of microbial genome sequences affords novel opportunities to re-examine the occurrence and distribution of nitrogen fixation genes. The current practice for computational prediction of nitrogen fixation is to use the presence of the nifH and/or nifD genes.ResultsBased on a careful comparison of the repertoire of nitrogen fixation genes in known diazotroph species we propose a new criterion for computational prediction of nitrogen fixation: the presence of a minimum set of six genes coding for structural and biosynthetic components, namely NifHDK and NifENB. Using this criterion, we conducted a comprehensive search in fully sequenced genomes and identified 149 diazotrophic species, including 82 known diazotrophs and 67 species not known to fix nitrogen. The taxonomic distribution of nitrogen fixation in Archaea was limited to the Euryarchaeota phylum; within the Bacteria domain we predict that nitrogen fixation occurs in 13 different phyla. Of these, seven phyla had not hitherto been known to contain species capable of nitrogen fixation. Our analyses also identified protein sequences that are similar to nitrogenase in organisms that do not meet the minimum-gene-set criteria. The existence of nitrogenase-like proteins lacking conserved co-factor ligands in both diazotrophs and non-diazotrophs suggests their potential for performing other, as yet unidentified, metabolic functions.ConclusionsOur predictions expand the known phylogenetic diversity of nitrogen fixation, and suggest that this trait may be much more common in nature than it is currently thought. The diverse phylogenetic distribution of nitrogenase-like proteins indicates potential new roles for anciently duplicated and divergent members of this group of enzymes.

Reconstruction and minimal gene requirements for the alternative iron-only nitrogenase in Escherichia coli

“…The inventory of genes required for diazotrophy varies greatly among diverse genera (23)(24)(25). A minimal core of three catalytic (nifHDK) and three biosynthetic (nifENB) genes is conserved in most diazotrophs (23,26), reflecting the core protein components required for FeMo-co biosynthesis in vitro (27).…”

“…A minimal core of three catalytic (nifHDK) and three biosynthetic (nifENB) genes is conserved in most diazotrophs (23,26), reflecting the core protein components required for FeMo-co biosynthesis in vitro (27). Other genes required for nitrogenase biosynthesis and activity are less well conserved or are substituted by other genes in the host.…”

“…Furthermore, some nif genes (e.g., electron transfer components) are specifically adapted to support nitrogenase activity in accordance with the physiological requirements of the host organism. Notably, alternative nitrogenases are only found in species that also encode a molybdenum-dependent nitrogenase (14,23).…”

“…In Rhodobacter capsulatus, diazotrophic growth dependent on the alternative FeFe nitrogenase is not influenced by mutations in either nifE or nifN, again indicating that biosynthesis of FeFe-co can take place in the absence of the NifEN scaffold (33). However, there are other nitrogenase-like sequences encoded in the R. capsulatus genome, which could potentially perform this function (23). In contrast, the increase of vnfENX expression in iron-only conditions in A. vinelandii (14), coupled with the absence of diazotrophic growth in double mutants lacking nifEN and vnfE (13), strongly supports a role for the VnfEN scaffold in FeFe nitrogenase maturation in this organism.…”

“…have been identified (42). However, the NifDK subunits in strains that lack nifEN or nifN represent a separate phylogenetic group, which appears to be associated with a thermophilic lifestyle (23). In Rhodobacter capsulatus, diazotrophic growth dependent on the alternative FeFe nitrogenase is not influenced by mutations in either nifE or nifN, again indicating that biosynthesis of FeFe-co can take place in the absence of the NifEN scaffold (33).…”