Nitrate reductases in Hydrogenobacter thermophilus with evolutionarily ancient features: distinctive localization and electron transfer.

Dissimilatory nitrate reductase (NAR) and assimilatory nitrate reductase (NAS) serve as key enzymes for nitrogen catabolism and anabolism in many organisms. We purified NAR and NAS from H. thermophilus, a hydrogen-oxidizing chemolithoautotroph belonging to the phylogenetically deepest branch in the Bacteria domain. Physiological contribution of these enzymes to nitrate respiration and assimilation was clarified by transcriptomic analysis and gene disruption experiments. These enzymes showed several features unreported in bacteria, such as the periplasmic orientation of NAR anchored with a putative transmembrane subunit and the specific electron transfer from a [4Fe-4S]-type ferredoxin to NAS. While some of their enzymatic properties are shared with NARs from archaea and with NASs from phototrophs, phylogenetic analysis indicated that H. thermophilus NAR and NAS have deep evolutionary origins that cannot be explained by a recent horizontal gene transfer event from archaea and phototrophs. These findings revealed the diversity of NAR and NAS in nonphotosynthetic bacteria, and they also implied that the outward orientation of NAR and the ferredoxin-dependent electron transfer of NAS are evolutionarily ancient features preserved in H. thermophilus.