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Bacterial pentaheme cytochrome c nitrite reductases (NrfAs) are key enzymes involved in the terminal step of dissimilatory nitrite reduction of the nitrogen cycle. Their structure and functions are well studied. Recently, a novel octaheme cytochrome c nitrite reductase (TvNiR) has been isolated from the haloalkaliphilic bacterium Thioalkalivibrio nitratireducens. Here we present high-resolution crystal structures of the apoenzyme and its complexes with the substrate (nitrite) and the inhibitor (azide). Both in the crystalline state and in solution, TvNiR exists as a stable hexamer containing 48 hemes-the largest number of hemes accommodated within one protein molecule known to date. The subunit of TvNiR consists of two domains. The N-terminal domain has a unique fold and contains three hemes. The catalytic C-terminal domain hosts the remaining five hemes, their arrangement, including the catalytic heme, being identical to that found in NrfAs. The complete set of eight hemes forms a spatial pattern characteristic of other multiheme proteins, including structurally characterized octaheme cytochromes. The catalytic machinery of TvNiR resembles that of NrfAs. It comprises the lysine residue at the proximal position of the catalytic heme, the catalytic triad of tyrosine, histidine, and arginine at the distal side, channels for the substrate and product transport with a characteristic gradient of electrostatic potential, and, finally, two conserved Ca(2+)-binding sites. However, TvNiR has a number of special structural features, including a covalent bond between the catalytic tyrosine and the adjacent cysteine and the unusual topography of the product channels that open into the void interior space of the protein hexamer. The role of these characteristic structural features in the catalysis by this enzyme is discussed.

Two strains of obligate chemolithoautotrophic sulfur-oxidizing bacteria were isolated from soda-lake sediments by enrichment culture with thiocyanate and nitrate at pH 9?9. The isolates were capable of growth with thiocyanate or thiosulfate as electron donor, either aerobically or anaerobically, and with nitrate or nitrite as electron acceptor. Cyanate was identified as an intermediate of thiocyanate oxidation, while sulfate, ammonia and dinitrogen gas were the final products. The anaerobic growth on thiocyanate plus nitrate was much slower (m max =0?006 h "1 ) than on thiosulfate plus nitrate (m max =0?02 h "1 ), while growth yields were similar (4?8 and 5?1 g protein mol "1 , respectively). On the basis of their phenotypic and genetic properties, strains ARhD 1 T and ARhD 2 are described as a novel species of the genus Thialkalivibrio, with the highest similarity to Thialkalivibrio denitrificans. The name Thialkalivibrio thiocyanodenitrificans sp. nov. is proposed for this novel species.

A novel moderately thermophilic, organotrophic, microaerophilic, facultatively chemolithotrophic bacterium, designated strain 506 T , was isolated from a deep-sea hydrothermal vent site at 13˚N in the East Pacific Rise. Cells were Gram-negative, non-motile rods. The organism grew in the temperature range 40-68˚C, with an optimum at 60˚C, and in the pH range 5?5-8?4, with an optimum around pH 7?5. The NaCl concentration for growth was in the range 10-50 g l 21 , with an optimum at 30 g l 21 . Strain 506 T grew chemoorganoheterotrophically with carbohydrates, proteinaceous substrates, organic acids and alcohols using oxygen or nitrate as electron acceptor. Alternatively, strain 506 T was able to grow lithoheterotrophically with molecular hydrogen as the energy source. The G+C content of the genomic DNA was 62?9 mol%. Phylogenetic analysis of the 16S rDNA sequence placed strain 506 T in the family Thermaceae. On the basis of phenotypic and phylogenetic data, strain 506 T (=DSM 14977 T =VKM B-2274 T ) is proposed as the type strain of a novel species in a new genus, Oceanithermus profundus gen. nov., sp. nov.

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