ANALYSIS OF CODON USAGE IN GENES FOR NITROGEN-FIXATION FROM PHYLOGENETICALLY DIVERSE DIAZOTROPHS

A cluster analysis based on codon usage in genes for biological nitrogen fixation (nif genes) grouped diazotrophs into three distinct classes: anaerobes, cyanobacteria, and aerobes. In the nif genes of Klebsiella pneumoniae there was no evidence for selection pressure in favor of highly translatable codons. However, in the nitrogen regulatory operon glnAntrBntrC of enteric bacteria the stoichiometrically high level of glutamine synthetase may be facilitated by the presence of efficiently translatable codons in glnA. The nif genes of the cyanobacterium Anabaena showed codon selection in favor of translational efficiency. Computation of codon adaptation indices for expression in heterologous systems indicated that the reading frames most suitable for expression of nif genes in Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae were present in azotobacters, clostridia, and cyanobacteria, respectively. In codon-usage-based cluster analysis, type 3 nitrogenase genes of Azotobacter vinelandii grouped along with the type 1 and type 2 genes. This is in contrast to the nucleotide sequence-based multiple alignment in which type 3 nitrogenase genes of A. vinelandii have been reported to cluster with entirely unrelated diazotrophs such as methanogens and clostridia. This may be indicative of lateral transfer of nif genes among widely divergent taxons. The chromosomal- and plasmid-located nif genes of rhizobia also cluster separately in nucleotide sequence-based analysis but showed similar codon usage. These analyses suggested that the phylogeny of nif genes drawn on the basis of nucleotide sequence homology was not masked by the taxon-specific pressure on codon usage.