The genus Thermococcus , comprised of sulfur-reducing hyperthermophilic archaea, belongs to the order Thermococcales in Euryarchaeota along with closely related Pyrococcus . members are ubiquitously present natural high-temperature environments, and therefore considered play a major role ecology metabolic activity microbial consortia within hot-water ecosystems. To obtain insight into this important genus, we have determined annotated complete 2,088,737-base genome kodakaraensis strain KOD1,...

A hyperthermophilic archaeal strain, KOD1, isolated from a solfatara on Kodakara Island, Japan, has previously been reported as Pyrococcus sp. KOD1. However, detailed phylogenetic tree, made possible by the recent accumulation of 16S rRNA sequences various species in order Thermococcales, indicated that strain KOD1 is member genus Thermococcus. We performed DNA-DNA hybridization tests against displayed high similarity terms ribosomal DNA sequences, including Thermococcus peptonophilus and...

N7-methylguanine at position 46 (m7G46) in tRNA is produced by methyltransferase (TrmB). To clarify the role of this modification, we made a trmB gene disruptant (ΔtrmB) Thermus thermophilus, an extreme thermophilic eubacterium. The absence TrmB activity cell extract from ΔtrmB strain and lack m7G46 modification tRNAPhe were confirmed enzyme assay, nucleoside analysis RNA sequencing. When was cultured high temperatures, several modified nucleotides hypo-modified addition to modification....

Pseudouridine at position 55 (Ψ55) in eubacterial tRNA is produced by TruB. To clarify the role of Ψ55 modification, we constructed a truB gene disruptant (Δ ) strain Thermus thermophilus which an extreme-thermophilic eubacterium. Unexpectedly, Δ exhibited severe growth retardation 50°C. We assumed that these phenomena might be caused lack RNA chaperone activity TruB, was previously hypothetically proposed others. confirm this idea, replaced genome with mutant genes, express TruB proteins...

Summary Myasthenia gravis (MG) is an autoimmune-mediated inflammatory disease of the neuromuscular junction. Previous studies animal MG models have suggested important roles cytokines in pathogenesis, but adequate on human are lacking. Using a multiplex suspension array system, we measured serum levels 27 cytokines/chemokines 47 anti-acetylcholine receptor antibody-positive patients with and 20 normal controls (NC) to investigate contribution toward pathogenesis. Correlations between...

Branching enzyme (BE) catalyzes formation of the branch points in glycogen and amylopectin by cleavage alpha-1,4 linkage its subsequent transfer to alpha-1,6 position. We have identified a novel BE encoded an uncharacterized open reading frame (TK1436) hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. TK1436 encodes conserved protein showing similarity members glycoside hydrolase family 57 (GH-57 family). At C terminus protein, two copies helix-hairpin-helix (HhH) motif were found....

We identified a novel regulator, Thermococcales glycolytic regulator (Tgr), functioning as both an activator and repressor of transcription in the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. Tgr (TK1769) displays similarity (28% identical) to Pyrococcus furiosus TrmB (PF1743), transcriptional regulating trehalose/maltose ATP-binding cassette transporter genes, but is more closely related (67%) paralog P. (PF0124). Growth tgr disruption strain (Deltatgr) displayed significant...

Hydrogenases catalyze the reversible oxidation of molecular hydrogen (H₂) and play a key role in energy metabolism microorganisms anaerobic environments. The hyperthermophilic archaeon Thermococcus kodakarensis KOD1, which assimilates organic carbon coupled with reduction elemental sulfur (S⁰) or H₂ generation, harbors three gene operons encoding [NiFe]-hydrogenase orthologs, namely, Hyh, Mbh, Mbx. In order to elucidate their functions vivo, disruption mutant for each ortholog was...

Significance The metal ions in proteins are correctly incorporated by specific metallochaperones. However, it remains unclear how metallochaperones regulate their binding affinity during acquisition of correct and deliver them to target proteins. In this study, we have determined the crystal structures a transient complex between Ni metallochaperone HypA its partner ATPase protein HypB AT , which incorporate ion into [NiFe] hydrogenase. reveal that induces conformational change through...

Fructose-1,6-bisphosphatase (FBPase) is one of the key enzymes gluconeogenic pathway. Although enzyme activity had been detected in Archaea, corresponding gene not identified until a presumable inositol monophosphatase from<i>Methanococcus jannaschii</i> was found to encode protein with both and FBPase activities. Here we display that from hyperthermophilic archaeon, <i>Thermococcus kodakaraensis</i> KOD1, which does correspond <i>M. jannaschii</i>, displays high activity. The strain KOD1...

Branching enzymes (BEs) catalyze the formation of branch points in glycogen and amylopectin by cleavage α-1,4 glycosidic bonds subsequent transfer to a new α-1,6 position. BEs generally belong glycoside hydrolase family 13 (GH13); however TK1436, isolated from hyperthermophilic archaeon Thermococcus kodakaraensis KOD1, is first GH57 member, which possesses BE activity. To date, only structure that had been determined GH13-type Escherichia coli. Herein, we have crystal TK1436 native state...

Long-chain and/or branched-chain polyamines are unique polycations found in thermophiles. Cytoplasmic were analyzed for cells cultivated at various growth temperatures the hyperthermophilic archaeon Thermococcus kodakarensis. Spermidine [34] and N4-aminopropylspermine [3(3)43] identified as major 60°C, amounts of increased temperature rose. To identify genes involved polyamine biosynthesis, a gene disruption study was performed. The open reading frames (ORFs) TK0240, TK0474, TK0882,...

Archaeal histones wrap DNA into complexes, designated archaeal nucleosomes, that resemble the tetrasome core of a eukaryotic nucleosome. Therefore, all interactions in vivo Thermococcus kodakarensis, most genetically versatile model species for research, must occur context histone-bound genome. Here we report construction and properties T. kodakarensis strains have TK1413 or TK2289 deleted, genes encode HTkA HTkB, respectively, two present this archaeon. All attempts to generate strain with...

We have previously determined the crystal structure of a novel pentagonal ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) from hyperthermophilic archaeon, Thermococcus kodakaraensis KOD1. Here we carried out biochemical studies to identify necessities and/or advantages this intriguing structure. The indicated presence three neighboring residues (Glu-63, Arg-66, and Asp-69), participating in ionic interactions within unique dimer-dimer interfaces. constructed single mutant proteins...

Background and purpose A single, oral dose of 3 mg/day tacrolimus, approved for myasthenia gravis ( MG ) treatment in Japan, was shown to reduce steroid anti‐acetylcholine receptor AC hR) antibody titers as well improve symptoms. However, no studies have investigated the association between tacrolimus concentration its clinical efficacy . In this study, we aimed determine optimal treatment. Methods The trough 51 patients with (positive anti‐ hR antibody, n = 48; negative anti‐muscle‐specific...

All archaeal genomes encode RNA polymerase (RNAP) subunits E and F that share a common ancestry with the eukaryotic RNAP A43 A14 (Pol I), Rpb7 Rpb4 II), C25 C17 III). By gene replacement, we have isolated mutants of Thermococcus kodakarensis subunit F-encoding (rpoF) deleted, but were unable to isolate lacking E-encoding (rpoE). Wild-type T. grows at temperatures ranging from 60 degrees C 100 C, optimally 85 DeltarpoF cells grew same rate as wild type 70 much slower lower cell densities C....

In eukaryotes, the replicative DNA helicase 'core' is minichromosome maintenance (Mcm) complex (MCM), forming a heterohexameric consisting of six subunits (Mcm2-7). Recent studies showed that CMG (Cdc45–MCM–GINS) actual body in replication fork progression complex. Archaea, Thermococcus kodakarensis harbors three genes encoding Mcm homologs on its genome, contrary to most archaea, which have only one homolog. It thus, high interest, whether and how these Mcms share their functions metabolism...

The hyperthermophilic archaeon Thermococcus kodakarensis can utilize sugars or pyruvate for growth. In the absence of elemental sulfur, electrons via oxidation these substrates are accepted by protons, generating molecular hydrogen (H2). hydrogenase responsible this reaction is a membrane-bound [NiFe]-hydrogenase (Mbh). study, we have examined several possibilities to increase protein levels Mbh in T. genetic engineering. Highest intracellular were achieved when promoter entire mbh operon...

A Gram-stain-negative, non-spore-forming, aerobic, oligotrophic bacterium (strain 262-7 T ) was isolated from a crack of white rock collected in the Skallen region Antarctica. Strain grew at temperatures between −4 and 30 °C, with optimal growth 25 °C. The pH range for 6.0 9.0, approximately 7.0. NaCl concentration allowing 0.0 1.0 %, an optimum 0.5 %. showed unprecedented morphological diversity response to conditions. Cells grown liquid medium were circular or ovoid smooth surfaces lag...

Significance The large subunit of [NiFe] hydrogenase undergoes Ni insertion into the active site by chaperone HypA during its posttranslational process (maturation process). However, detailed mechanism has been elusive. Here, we report structures an immature HyhL in complex with HypA, which enable us to infer a series conformational changes involved delivery. show that N-terminal tail interacts Ni-binding domain HypA. Structural comparison mature one reveals may induce spatial rearrangement...