The polyextremophile Natranaerobius thermophilus adopts a dual adaptive strategy to long-term salinity stress, simultaneously accumulating compatible solutes and K.

The bacterium Natranaerobius thermophilus is an extremely halophilic alkalithermophile that can thrive under conditions of high salinity (3.3-3.9 M Na+), alkaline pH (9.5), and elevated temperature (53°C). To understand the molecular mechanisms of salt adaptation in N. thermophilus, it is essential to investigate the protein, mRNA, and key metabolite levels on a molecular basis. Based on proteome profiling of N. thermophilus under 3.1, 3.7, and 4.3 M Na+ conditions compared to 2.5 M Na+ condition, we discovered that a hybrid strategy, combining the "compatible solute" and "salt-in" mechanisms, was utilized for osmotic adjustment dur ing the long-term salinity adaptation of N. thermophilus. The mRNA level of key proteins and the intracellular content of compatible solutes and K+ support this conclusion. Specifically, N. thermophilus employs the glycine betaine ABC transporters (Opu and ProU families), Na+/solute symporters (SSS family), and glutamate and proline synthesis pathways to adapt to high salinity. The intracellular content of compatible solutes, including glycine betaine, glutamate, and proline, increases with rising salinity levels in N. thermophilus. Additionally, the upregulation of Na+/ K+/ H+ transporters facilitates the maintenance of intracellular K+ concentration, ensuring cellular ion homeostasis under varying salinities. Furthermore, N. thermophilus exhibits cytoplasmic acidification in response to high Na+ concentrations. The median isoelectric points of the upregulated proteins decrease with increasing salinity. Amino acid metabolism, carbohydrate and energy metabolism, membrane transport, and bacterial chemotaxis activities contribute to the adaptability of N. thermophilus under high salt stress. This study provides new data that support further elucidating the complex adaptation mechanisms of N. thermophilus under multiple extremes.IMPORTANCEThis study represents the first report of simultaneous utilization of two salt adaptation mechanisms within the Clostridia class in response to long-term salinity stress.

Reclassification of the genus Natronolimnobius: proposal of two new genera, Natronolimnohabitans gen. nov. to accommodate Natronolimnobius innermongolicus and Natrarchaeobaculum gen. nov. to accommodate Natronolimnobius aegyptiacus and Natronolimnobius sulfurireducens.

The genus Natronolimnobius, currently including four species, is a member of the order Natrialbales, class Halobacteria, and consists of obligately alkaliphilic and extremely halophilic members found exclusively in highly alkaline hypersaline soda lakes. The species were classified into this genus mostly based on phylogenetic analysis of the 16S rRNA gene. However, a more advanced phylogenomic reconstruction based on 122 conserved single-copy archaeal protein markers clearly indicates a polyphyletic origin of the species included into this genus, thus warranting its reclassification into three separate genera. We therefore propose to transfer Nlb. innermongolicus (type strain N-1311) to a new genus Natronolimnohabitans as Nlh. innermongolicus comb. nov. and to transfer Nlb. aegyptiacus (type strain JW/NM-HA 15) and Nlb. sulfurireducens (type strain AArc1) to a new genus Natrarchaeobaculum as Nbl. aegyptiacum comb. nov. and Nbl. sulfurireducens comb. nov. The phylogenomic differentiation of these four species is also supported by the ANI/AAI distances and unique phenotypes. The most important physiological differences includes a previously unreported ability for cellulose and xylan utilization in Nlb. baerhuensis, thermophily in Nbl. aegyptiacus and anaerobic sulfur respiration in Nbl. sulfurireducens. We further present an emended description of Natronolimnobius baerhuensis.

Natronolimnobius aegyptiacus sp. nov., an extremely halophilic alkalithermophilic archaeon isolated from the athalassohaline Wadi An Natrun, Egypt.

An obligately aerobic extremely halophilic alkalithermophilic archaeon, strain JW/NM-HA 15T, was isolated from the sediments of Wadi An Natrun in Egypt. Phylogenetic analysis based on 16S rRNA and rpoB' gene sequences indicated that it belongs to the family Natrialbaceae of the order Natrialbales. The closest relatives were Natronolimnobius baerhuensis IHC-005T and Natronolimnobius innermongolicus N-1311T (95.3 and 94.5 % 16S rRNA gene sequence similarity, respectively). Genome relatedness between strain JW/NM-HA 15T and its neighbours was evaluated using average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity with the values of 75.7-85.0, 18.1-20.0, and 70.2-71.0%, respectively. Cells were obligately aerobic, rod-shaped, non-motile, Gram-stain-negative and chemo-organotrophic. The strain grew in the presence of 2.57 M to saturating Na+ (optimum 3.25-4.60 M Na+), at pH55 °C 7.5-10.5 (optimum pH55 °C 9.0-9.5), and at 30-56 °C (optimum 52 °C). The major polar lipids consisted of phosphatidylglycerol, methylated phosphatidylglycerolphosphate and two phospholipids. The complete genome size of strain JW/NM-HA 15T is approximately 3.93 Mb, with a DNA G+C content of 64.1 mol%. On the basis of phylogenetic features, genomic relatedness, phenotypic and chemotaxonomic data, strain JW/NM-HA 15T was thus considered to represent a novel species within the genus Natronolimnobius, for which the name Natronolimnobius aegyptiacus sp. nov. is proposed. The type strain is JW/NM-HA 15T (=ATCC BAA-2088T =DSM 23470T).

Biochemical characterization of halophilic, alkalithermophilic amylopullulanase PulD7 and truncated amylopullulanases PulD7ΔN and PulD7ΔC.

A pullulanase, PulD7, was identified in the genome of the halophilic, alkalithermophilic isolate Alkalilimnicola sp. NM-DCM-1. PulD7 is 701 amino acids large with a carbohydrate binding module (CBM) 48 at the N-terminal. The full length PulD7 and N- and C-terminal truncated versions were cloned, heterologously expressed and functionally characterized. PulD7 displayed maximal activity at 55 °C, pH 9.5 and 2 M NaCl. PulD7 had good thermal stability, with a half-life of 693 min at 50 °C. PulD7 is an amylopullulanase, hydrolyzing both α­1,4­ and α­1,6­glycosidic bonds in soluble starch and pullulan, respectively. PulD7 was resistant to chemical reagents, including organic solvents (dimethyl sulfoxide, methanol, benzene, 20% v/v), reducing agents (ß-mercaptoethanol, 5% v/v), surfactants (SDS and Tween 20, 5% v/v), the divalent chelator ethylene diamine tetra acetic acid (5 mM), and the chemical denaturant urea (8 M). PulD7 was not calcium-dependent. PulD7 was able to bind raw starch granules, reaching 52% binding in 3 h. The N-and C-terminal truncated forms of PulD7 had similar biochemical characteristics. PulD7ΔC had higher specific activity and halotolerance. The N-terminally truncated PulD7ΔN hydrolyzed amylose only, indicating that CBM48 is essential for binding branched substrates. PulD7 has unique characteristics giving it strong potential for application in biotechnological industries.

Biochemical characterization of a halophilic, alkalithermophilic protease from Alkalibacillus sp. NM-Da2.

An extracellular, halophilic, alkalithermophilic serine protease from the halo-alkaliphilic Alkalibacillus sp. NM-Da2 was purified to homogeneity by ethanol precipitation and anion-exchange chromatography. The purified protease was a monomeric enzyme with an approximate molecular mass of 35 kDa and exhibited maximal activity at 2.7 M NaCl, pH55 °C 9 and 56 °C. The protease showed great temperature stability, retaining greater than 80 % of initial activity after 2 h incubation at 55 °C. The protease was also extremely pH tolerant, retaining 80 % of initial activity at pH55 °C 10.5 after 30 min incubation. Protease hydrolyzed complex substrates, displaying activity on yeast extract, tryptone, casein, gelatin and peptone. Protease activity was inhibited at casein concentrations greater than 1.2 mg/mL. The enzyme was stable and active in 40 % (v/v) solutions of isopropanol, ethanol and benzene and was stable in the presence of the polysorbate surfactant Tween 80. Activity was stimulated with the oxidizing agent hydrogen peroxide. Inhibition with phenyl methylsulfonylfluoride indicates it is a serine protease. Synthetic saline wastewater treated with the protease showed 50 % protein removal after 5 h. Being halophilic, alkaliphilic and thermophilic, in addition to being resistant to organic solvents, this protease has potential for various applications in biotechnological and pharmaceutical industries.

Distinct structural features of Rex-family repressors to sense redox levels in anaerobes and aerobes.

The Rex-family repressors sense redox levels by alternative binding to NADH or NAD(+). Unlike other Rex proteins that regulate aerobic respiration, RSP controls ethanol fermentation in the obligate anaerobe Thermoanaerobacter ethanolicus JW200(T). It is also found in other anaerobic microorganisms. Here we present the crystal structures of apo-RSP, RSP/NADH and RSP/NAD(+)/DNA, which are the first structures of Rex-family members from an obligate anaerobe. RSP functions as a homodimer. It assumes an open conformation when bound to the operator DNA and a closed conformation when not DNA-bound. The DNA binds to the N-terminal winged-helix domain and the dinucleotide, either reduced or oxidized, binds to the C-terminal Rossmann-fold domain. The two distinct orientations of nicotinamide ring, anti in NADH and syn in NAD(+), give rise to two sets of protein-ligand interactions. Consequently, NADH binding makes RSP into a closed conformation, which does not bind to DNA. Both the conserved residues and the DNA specificity of RSP show a number of variations from those of the aerobic Rex, reflecting different structural bases for redox-sensing by the anaerobic and aerobic Rex-family members.

Anaerobic thermophiles.

The term "extremophile" was introduced to describe any organism capable of living and growing under extreme conditions. With the further development of studies on microbial ecology and taxonomy, a variety of "extreme" environments have been found and an increasing number of extremophiles are being described. Extremophiles have also been investigated as far as regarding the search for life on other planets and even evaluating the hypothesis that life on Earth originally came from space. The first extreme environments to be largely investigated were those characterized by elevated temperatures. The naturally "hot environments" on Earth range from solar heated surface soils and water with temperatures up to 65 °C, subterranean sites such as oil reserves and terrestrial geothermal with temperatures ranging from slightly above ambient to above 100 °C, to submarine hydrothermal systems with temperatures exceeding 300 °C. There are also human-made environments with elevated temperatures such as compost piles, slag heaps, industrial processes and water heaters. Thermophilic anaerobic microorganisms have been known for a long time, but scientists have often resisted the belief that some organisms do not only survive at high temperatures, but actually thrive under those hot conditions. They are perhaps one of the most interesting varieties of extremophilic organisms. These microorganisms can thrive at temperatures over 50 °C and, based on their optimal temperature, anaerobic thermophiles can be subdivided into three main groups: thermophiles with an optimal temperature between 50 °C and 64 °C and a maximum at 70 °C, extreme thermophiles with an optimal temperature between 65 °C and 80 °C, and finally hyperthermophiles with an optimal temperature above 80 °C and a maximum above 90 °C. The finding of novel extremely thermophilic and hyperthermophilic anaerobic bacteria in recent years, and the fact that a large fraction of them belong to the Archaea has definitely made this area of investigation more exciting. Particularly fascinating are their structural and physiological features allowing them to withstand extremely selective environmental conditions. These properties are often due to specific biomolecules (DNA, lipids, enzymes, osmolites, etc.) that have been studied for years as novel sources for biotechnological applications. In some cases (DNA-polymerase, thermostable enzymes), the search and applications successful exceeded preliminary expectations, but certainly further exploitations are still needed.

Halophilic alkali- and thermostable amylase from a novel polyextremophilic Amphibacillus sp. NM-Ra2.

Extracellular gluco-amylo-pullulanase from Amphibacillus sp. NM-Ra2 was purified to homogeneity by ethanol precipitation, anion exchange chromatography and gel filtration chromatography. Molecular mass of the enzyme was 50kDa (SDS-PAGE). The enzyme showed maximal activity at 1.9 M NaCl, pH50°C 8.0 and 54°C and was active from 0 to 4.3 M NaCl and 37 to 65°C. The enzyme was inhibited by EDTA and was stable and active in the presence of PMSF, DTT, H2O2, Triton-X-100, Tween 20 and Tween 80. Ca2+ is inessential for activity. The amylase was stimulated with K+ and inhibited with Cu2+ and Mg2+. Hg2+, Zn2+ and Fe2+ had no effect on activity. Amylase was stable and active in the presence of ethanol, methanol and benzene (25%, v/v). The enzyme hydrolyzed linear and branched polysaccharides including pullulan, glycogen and amylopectin, and hydrolyzed raw wheat starch and raw corn starch (14.6% and 13.5% over 2 h). Amylase activity was inhibited by soluble starch concentrations greater than 0.3%. The major products of soluble starch hydrolysis were maltose and maltotriose. The amylase, being halophilic and alkali-thermostable, in addition to being resistant to surfactants, oxidizing agents and organic solvents, can find applications in the starch processing, pharmaceutical, food and paper/pulp industries.

Draft Genome Sequence of the Fast-Growing Bacterium Vibrio natriegens Strain DSMZ 759.

Vibrio natriegens is a Gram-negative bacterium known for its extremely short doubling time. Here we present the annotated draft genome sequence of Vibrio natriegens strain DSMZ 759, with the aim of providing insights about its high growth rate.

Multilocus sequence analysis of Thermoanaerobacter isolates reveals recombining, but differentiated, populations from geothermal springs of the Uzon Caldera, Kamchatka, Russia.

Thermal environments have island-like characteristics and provide a unique opportunity to study population structure and diversity patterns of microbial taxa inhabiting these sites. Strains having ≥98% 16S rRNA gene sequence similarity to the obligately anaerobic Firmicutes Thermoanaerobacter uzonensis were isolated from seven geothermal springs, separated by up to 1600 m, within the Uzon Caldera (Kamchatka, Russian Far East). The intraspecies variation and spatial patterns of diversity for this taxon were assessed by multilocus sequence analysis (MLSA) of 106 strains. Analysis of eight protein-coding loci (gyrB, lepA, leuS, pyrG, recA, recG, rplB, and rpoB) revealed that all loci were polymorphic and that nucleotide substitutions were mostly synonymous. There were 148 variable nucleotide sites across 8003 bp concatenates of the protein-coding loci. While pairwise F ST values indicated a small but significant level of genetic differentiation between most subpopulations, there was a negligible relationship between genetic divergence and spatial separation. Strains with the same allelic profile were only isolated from the same hot spring, occasionally from consecutive years, and single locus variant (SLV) sequence types were usually derived from the same spring. While recombination occurred, there was an "epidemic" population structure in which a particular T. uzonensis sequence type rose in frequency relative to the rest of the population. These results demonstrate spatial diversity patterns for an anaerobic bacterial species in a relative small geographic location and reinforce the view that terrestrial geothermal springs are excellent places to look for biogeographic diversity patterns regardless of the involved distances.

Structural and functional analyses of catalytic domain of GH10 xylanase from Thermoanaerobacterium saccharolyticum JW/SL-YS485.

Xylanases are capable of decomposing xylans, the major components in plant cell wall, and releasing the constituent sugars for further applications. Because xylanase is widely used in various manufacturing processes, high specific activity, and thermostability are desirable. Here, the wild-type and mutant (E146A and E251A) catalytic domain of xylanase from Thermoanaerobacterium saccharolyticum JW/SL-YS485 (TsXylA) were expressed in Escherichia coli and purified subsequently. The recombinant protein showed optimal temperature and pH of 75°C and 6.5, respectively, and it remained fully active even after heat treatment at 75°C for 1 h. Furthermore, the crystal structures of apo-form wild-type TsXylA and the xylobiose-, xylotriose-, and xylotetraose-bound E146A and E251A mutants were solved by X-ray diffraction to high resolution (1.32-1.66 Å). The protein forms a classic (ß/α)8 folding of typical GH10 xylanases. The ligands in substrate-binding groove as well as the interactions between sugars and active-site residues were clearly elucidated by analyzing the complex structures. According to the structural analyses, TsXylA utilizes a double displacement catalytic machinery to carry out the enzymatic reactions. In conclusion, TsXylA is effective under industrially favored conditions, and our findings provide fundamental knowledge which may contribute to further enhancement of the enzyme performance through molecular engineering.

The substrate/product-binding modes of a novel GH120 ß-xylosidase (XylC) from Thermoanaerobacterium saccharolyticum JW/SL-YS485.

Xylan-1,4-ß-xylosidase (ß-xylosidase) hydrolyses xylo-oligomers at their non-reducing ends into individual xylose units. Recently, XylC, a ß-xylosidase from Thermoanaerobacterium saccharolyticum JW/SL-YS485, was found to be structurally different from corresponding glycosyl hydrolases in the CAZy database (http://www.cazy.org/), and was subsequently classified as the first member of a novel family of glycoside hydrolases (GH120). In the present paper, we report three crystal structures of XylC in complex with Tris, xylobiose and xylose at 1.48-2.05 Å (1 Å=0.1 nm) resolution. XylC assembles into a tetramer, and each monomer comprises two distinct domains. The core domain is a right-handed parallel ß-helix (residues 1-75 and 201-638) and the flanking region (residues 76-200) folds into a ß-sandwich domain. The enzyme contains an open carbohydrate-binding cleft, allowing accommodation of longer xylo-oligosaccharides. On the basis of the crystal structures and in agreement with previous kinetic data, we propose that XylC cleaves the glycosidic bond by the retaining mechanism using two acidic residues Asp382 (nucleophile) and Glu405 (general acid/base). In addition to the active site, nine other xylose-binding sites were consistently observed in each of the four monomers, providing a possible reason for the high tolerance of product inhibition.

Crystallization and preliminary X-ray diffraction analysis of a novel GH120 ß-xylosidase (XylC) from Thermoanaerobacterium saccharolyticum JW/SL-YS485.

Xylosidases hydrolyze xylopolymers at the nonreducing end to free xylose units. The ß-xylosidase (XylC) from Thermoanaerobacterium saccharolyticum JW/SL-YS485 was expressed in Escherichia coli and the recombinant protein was purified and crystallized. A BLASTP search with the XylC protein sequence showed that no similar structure had previously been solved. XylC was classified as a member of the new glycoside hydrolase family GH120 according to the CAZy website (http://www.cazy.org/). Crystals belonging to the monoclinic space group P2(1), with unit-cell parameters a = 88.36, b = 202.20, c = 99.87 Å, ß = 99.04°, were obtained by the sitting-drop vapour-diffusion method and diffracted to 2.2 Šresolution. Structure determination using MIR and MAD methods is in progress.

Life under multiple extreme conditions: diversity and physiology of the halophilic alkalithermophiles.

Around the world, there are numerous alkaline, hypersaline environments that are heated either geothermally or through intense solar radiation. It was once thought that such harsh environments were inhospitable and incapable of supporting a variety of life. However, numerous culture-dependent and -independent studies revealed the presence of an extensive diversity of aerobic and anaerobic bacteria and archaea that survive and grow under these multiple harsh conditions. This diversity includes the halophilic alkalithermophiles, a novel group of polyextremophiles that require for growth and proliferation the multiple extremes of high salinity, alkaline pH, and elevated temperature. Life under these conditions undoubtedly involves the development of unique physiological characteristics, phenotypic properties, and adaptive mechanisms that enable control of membrane permeability, control of intracellular osmotic balance, and stability of the cell wall, intracellular proteins, and other cellular constituents. This minireview highlights the ecology and growth characteristics of the extremely halophilic alkalithermophiles that have been isolated thus far. Biochemical, metabolic, and physiological properties of the extremely halophilic alkalithermophiles are described, and their roles in resistance to the combined stressors of high salinity, alkaline pH, and high temperature are discussed. The isolation of halophilic alkalithermophiles broadens the physicochemical boundaries for life and extends the boundaries for the combinations of the maximum salinity, pH, and temperature that can support microbial growth.

Comparative geochemical and microbiological characterization of two thermal pools in the Uzon Caldera, Kamchatka, Russia.

Arkashin Schurf (Arkashin) and Zavarzin Spring (Zavarzin), two active thermal pools in the Uzon Caldera, Kamchatka, Russia, were studied for geochemical and microbiological characterization. Arkashin, the smaller of the two pools, had broader temperature and pH ranges, and the sediments had higher concentrations of total As (4,250 mg/kg) relative to Zavarzin (48.9 mg/kg). Glycerol dialkyl glycerol tetraether profiles represented distinct archaeal communities in each pool and agreed well with previous studies of these pools. Although no archaeal 16S rRNA sequences were recovered from Arkashin, sequences recovered from Zavarzin were mostly representatives of the Crenarchaeota and "Korarchaeota," and 13% of the sequences were unclassifiable. The bacterial community in Arkashin was dominated by uncultured "Bacteroidetes," Hydrogenobaculum of the Aquificales and Variovorax of the Betaproteobacteria, and 19% of the sequences remained unclassified. These results were consistent with other studies of As-rich features. The most abundant members of the Zavarzin bacterial community included the Chloroflexi, as well as members of the classes Deltaproteobacteria and Clostridia. In addition, 24% of the sequences were unclassified and at least 5% of those represent new groups among the established Bacterial phyla. Ecological structure in each pool was inferred from taxonomic classifications and bulk stable isotope δ values of C, N, and S. Hydrogenobaculum was responsible for primary production in Arkashin. However, in Zavarzin, the carbon source appeared to be allochthonous to the identified bacterial community members. Additionally, sequences related to organisms expected to participate in N and S cycles were identified from both pools.

Amphibacillus cookii sp. nov., a facultatively aerobic, spore-forming, moderately halophilic, alkalithermotolerant bacterium.

Novel strains of facultatively aerobic, moderately alkaliphilic and facultatively halophilic bacteria were isolated from a sediment sample taken from the Southern Arm of Great Salt Lake, Utah. Cells of strain JW/BP-GSL-QD(T) (and related strains JW/BP-GSL-RA and JW/BP-GSL-WB) were rod-shaped, spore-forming, motile bacteria with variable Gram-staining. Strain JW/BP-GSL-QD(T) grew under aerobic conditions between 14.5 and 47 °C (optimum 39 °C), in the pH(37 °C) range 6.5-10.3 (optimum pH(37 °C) 8.0), and between 0.1 and 4.5 M Na(+) (optimum 0.9 M Na(+)). No growth was observed in the absence of supplemented Na(+). Strain JW/BP-GSL-QD(T) utilized L-arabinose, D-fructose, D-galactose, D-glucose, inulin, lactose, maltose, mannitol, D-mannose, pyruvate, D-ribose, D-sorbitol, starch, trehalose, xylitol and D-xylose under both aerobic and anaerobic conditions, and used ethanol and methanol only under aerobic conditions. Strains JW/BP-GSL-WB and JW/BP-GSL-RA had the same profiles except that methanol was not used aerobically. During growth on glucose, the major organic compounds formed under aerobic conditions were acetate and lactate, and under anaerobic conditions, the fermentation products were formate, acetate, lactate and ethanol. Oxidase and catalase activities were not detected and cytochrome was absent. No respiratory quinones were detected. The main cellular fatty acids were iso-C(15 : 0) (39.1 %) and anteiso-C(15 : 0) (36.3 %). Predominant polar lipids were diphosphatidylglycerol, phosphatidylglycerol and an unknown phospholipid. Additionally, a small amount of an unknown glycolipid was detected. The DNA G+C content of strain JW/BP-GSL-QD(T) was 35.4 mol% (determined by HPLC). For strain JW/BP-GSL-QD(T) the highest degree of 16S rRNA gene sequence similarity was found with Amphibacillus jilinensis (98.6 %), Amphibacillus sediminis (96.7 %) and Amphibacillus tropicus (95.6 %). The level of DNA-DNA relatedness between strain JW/BP-GSL-QD(T) and A. jilinensis Y1(T) was 58 %. On the basis of physiological, chemotaxonomic and phylogenetic data, strain JW/BP-GSL-QD(T) represents a novel species of the genus Amphibacillus, for which the name Amphibacillus cookii sp. nov. is proposed. The type strain is JW/BP-GSL-QD(T) (= ATCC BAA-2118(T) = DSM 23721(T)).

Correlation of genomic and physiological traits of thermoanaerobacter species with biofuel yields.

Thermophilic anaerobic noncellulolytic Thermoanaerobacter species are of great biotechnological importance in cellulosic ethanol production due to their ability to produce high ethanol yields by simultaneous fermentation of hexose and pentose. Understanding the genome structure of these species is critical to improving and implementing these bacteria for possible biotechnological use in consolidated bioprocessing schemes (CBP) for cellulosic ethanol production. Here we describe a comparative genome analysis of two ethanologenic bacteria, Thermoanaerobacter sp. X514 and Thermoanaerobacter pseudethanolicus 39E. Compared to 39E, X514 has several unique key characteristics important to cellulosic biotechnology, including additional alcohol dehydrogenases and xylose transporters, modifications to pentose metabolism, and a complete vitamin B12 biosynthesis pathway. Experimental results from growth, metabolic flux, and microarray gene expression analyses support genome sequencing-based predictions which help to explain the distinct differences in ethanol production between these strains. The availability of whole-genome sequence and comparative genomic analyses will aid in engineering and optimizing Thermoanaerobacter strains for viable CBP strategies.

Complete genome sequence of the anaerobic, halophilic alkalithermophile Natranaerobius thermophilus JW/NM-WN-LF.

The genome of the anaerobic halophilic alkalithermophile Natranaerobius thermophilus consists of one 3,165,557-bp chromosome and two plasmids (17,207 bp and 8,689 bp). The present study is the first to report the completely sequenced genome of an anaerobic polyextremophile and genes associated with roles in regulation of intracellular osmotic pressure, pH homeostasis, and growth at elevated temperatures.