Assessing Horizontal Transfer of nifHDK Genes in Eubacteria: Nucleotide Sequence of nifK

The structural genes for nitrogenase, nifK, nifD, and nifr-r, are crucial for nitrogen fixation. Previous phylogenetic analysis of the amino acid sequence of nifH suggested that this gene had been horizontally transferred from a proteobacterium to the gram-positive/cyanobacterial clade, although the confounding effects of paralogous comparisons made interpretation of the data difficult. An additional test of nifgene horizontal transfer using nzjD was made, but the NifD phylogeny lacked resolution. Here nifgene phylogeny is addressed with a phylogenetic analysis of a third and longer nifgene, nzyK. As part of the study, the nifK gene of the key taxon Frankia was sequenced. Parsimony and some distance analyses of the nifK amino acid sequences provide support for vertical descent of nzyK, but other distance trees provide support for the lateral transfer of the gene. Bootstrap support was found for both hypotheses in all trees; the nifK data do not definitively favor one or the other hypothesis. A parsimony analysis of NifH provides support for horizontal transfer in accord with previous reports, although bootstrap analysis also shows some support for vertical descent of the orthologous nifH genes. A wider sampling of taxa and more sophisticated methods of phylogenetic inference are needed to understand the evolution of nifgenes. The nifgenes may also be powerful phylogenetic tools. If nifK evolved by vertical descent, it provides strong evidence that the cyanobacteria and proteobacteria are sister groups to the exclusion of the firmicutes, whereas 16s rRNA sequences are unable to resolve the relationships of these three major eubacterial lineages.