Identification of Initial Colonizing Bacteria in Dental Plaques from Young Adults Using Full-Length 16S rRNA Gene Sequencing.

Development of dental plaque begins with the adhesion of salivary bacteria to the acquired pellicle covering the tooth surface. In this study, we collected dental plaque formed on hydroxyapatite disks for 6 h from 74 young adults and identified initial colonizing taxa based on full-length 16S rRNA gene sequences. A long-read, single-molecule sequencer, PacBio Sequel, provided 100,109 high-quality full-length 16S rRNA gene sequence reads from the early plaque microbiota, which were assigned to 90 oral bacterial taxa.

Characteristics of the Salivary Microbiota in Patients With Various Digestive Tract Cancers.

The salivary microbiota is constantly swallowed and delivered to the digestive tract. These bacteria may be associated with gastrointestinal diseases. This case-control study examined the salivary microbiota in patients with digestive tract cancer (DTC) and evaluated their differential distribution based on the cancer sites.

Effect of coffee on the compositional shift of oral indigenous microbiota cultured in vitro.

Coffee is a widely consumed beverage containing organic compounds with antibacterial activity. To investigate its possible effect on the growth of oral indigenous microbiota, saliva samples collected from nine young adults were inoculated into brain heart infusion (BHI) medium with or without addition of coffee compounds and cultured at 37°C in 5% CO for 12 h. The total bacterial density and composition after cultivation for 0, 6, and 12 h were determined by quantitative PCR analysis and 16S rRNA gene sequencing, respectively.

Characterization of oral microbiota and acetaldehyde production.

: has been reported to be a high producer of acetaldehyde (ACH), a carcinogen, from ethanol , but no information exists regarding whether the ACH production depends on oral microbiota profiles. To explore the salivary microbiota profiles with respect to ACH production ability in the oral cavity using a cross-sectional design. Using 16S rRNA gene amplicon sequencing, we classified 100 saliva samples into two types of communities (I and II).

Genome Replication in Independent of Cdc6 and an Origin of Replication.

The initiation of DNA replication is typically tightly regulated by proteins that form initiation complexes at specific sequences known as replication origins. In Archaea and Eukaryotes, Cdc6, a near-universally conserved protein binds and facilitates the origin-dependent assembly of the replicative apparatus. TK1901 encodes Cdc6 in but, as we report here, TK1901 and the presumed origin of replication can be deleted from the genome of this hyperthermophilic Archaeon without any detectable effects on growth, genetic competence or the ability to support autonomous plasmid replication.

Crystal structures of the carbamoylated and cyanated forms of HypE for [NiFe] hydrogenase maturation.

Hydrogenase pleiotropically acting protein (Hyp)E plays a role in biosynthesis of the cyano groups for the NiFe(CN)2CO center of [NiFe] hydrogenases by catalyzing the ATP-dependent dehydration of the carbamoylated C-terminal cysteine of HypE to thiocyanate. Although structures of HypE proteins have been determined, until now there has been no structural evidence to explain how HypE dehydrates thiocarboxamide into thiocyanate. Here, we report the crystal structures of the carbamoylated and cyanated forms of HypE from Thermococcus kodakarensis in complex with nucleotides at 1.

Identification and structure of a novel archaeal HypB for [NiFe] hydrogenase maturation.

HypB (metal-binding GTPase) and HypA (nickel metallochaperone) are required for nickel insertion into [NiFe] hydrogenase. However, the HypB homolog proteins are not found in some archaeal species including Thermococcales. In this article, we identify a novel archaeal Mrp/MinD family ATPase-type HypB from Thermococcus kodakarensis (Tk-mmHypB) and determine its crystal structure.

Crystal structures of the HypCD complex and the HypCDE ternary complex: transient intermediate complexes during [NiFe] hydrogenase maturation.

[NiFe] hydrogenase maturation represents one of the most dynamic and sophisticated processes in metallocenter assembly. The Fe(CN)(2)CO moiety of [NiFe] hydrogenases is assembled via unknown transient interactions among specific maturation proteins HypC (metallochaperone), HypD (redox protein), and HypE (cyanide synthesis/donor). Here, we report the structures of the HypC-HypD and HypC-HypD-HypE complexes, providing a view of the transient interactions that take place during the maturation process.

Structure of the [NiFe]-hydrogenase maturation protein HypF from Thermococcus kodakarensis KOD1.

HypF is involved in the biosynthesis of the CN ligand of the NiFe(CN)(2)CO centre of [NiFe]-hydrogenases. Here, the full-length structure of HypF from Thermococcus kodakarenesis is reported at 4.5 Å resolution.

Histone and TK0471/TrmBL2 form a novel heterogeneous genome architecture in the hyperthermophilic archaeon Thermococcus kodakarensis.

Being distinct from bacteria and eukaryotes, Archaea constitute a third domain of living things. The DNA replication, transcription, and translation machineries of Archaea are more similar to those of eukaryotes, whereas the genes involved in metabolic processes show more similarity to their bacterial counterparts. We report here that TK0471/TrmB-like 2 (TrmBL2), in addition to histone, is a novel type of abundant chromosomal protein in the model euryarchaeon Thermococcus kodakarensis .

Isoprenoid biosynthesis in Archaea--biochemical and evolutionary implications.

Isoprenoids are indispensable for all types of cellular life in the Archaea, Bacteria, and Eucarya. These membrane-associated molecules are involved in a wide variety of vital biological functions, ranging from compartmentalization and stability, to protection and energy-transduction. In Archaea, isoprenoid compounds constitute the hydrophobic moiety of the typical ether-linked membrane lipids.

Formate-driven growth coupled with H(2) production.

Although a common reaction in anaerobic environments, the conversion of formate and water to bicarbonate and H(2) (with a change in Gibbs free energy of ΔG° = +1.3 kJ mol(-1)) has not been considered energetic enough to support growth of microorganisms. Recently, experimental evidence for growth on formate was reported for syntrophic communities of Moorella sp.

Thermococcus kodakarensis mutants deficient in di-myo-inositol phosphate use aspartate to cope with heat stress.

Many of the marine microorganisms which are adapted to grow at temperatures above 80 degrees C accumulate di-myo-inositol phosphate (DIP) in response to heat stress. This led to the hypothesis that the solute plays a role in thermoprotection, but there is a lack of definitive experimental evidence. Mutant strains of Thermococcus kodakarensis (formerly Thermococcus kodakaraensis), manipulated in their ability to synthesize DIP, were constructed and used to investigate the involvement of DIP in thermoadaptation of this archaeon.

Crystal structure of HypA, a nickel-binding metallochaperone for [NiFe] hydrogenase maturation.

HypA is one of the auxiliary proteins involved in the maturation of [NiFe] hydrogenases. By an unknown mechanism, HypA functions as a metallochaperone in the insertion of the Ni atom into hydrogenases. We have determined the crystal structures of HypA from Thermococcus kodakaraensis KOD1 in both monomeric and dimeric states.

Polarity in archaeal operon transcription in Thermococcus kodakaraensis.

An in vivo archaeal gene reporter system has been established based on TK1761, a gene that encodes a nonessential beta-glycosidase in Thermococcus kodakaraensis. Following the introduction of nonsense codons into promoter-proximal genes, polarity in operon expression in this archaeon has been established by both microarray hybridization assays and a reporter gene expression system.

Crystallization and preliminary X-ray crystallographic study of [NiFe]-hydrogenase maturation factor HypE from Thermococcus kodakaraensis KOD1.

The hydrogenase maturation protein HypE is involved in the biosynthesis of the CN ligands of the active-site iron of [NiFe] hydrogenases using carbamoylphosphate as a substrate. Here, the crystallization and preliminary crystallographic analysis of HypE from Thermococcus kodakaraensis KOD1 are reported. Crystals of HypE (338 amino acids, 35.

Crystal structures of [NiFe] hydrogenase maturation proteins HypC, HypD, and HypE: insights into cyanation reaction by thiol redox signaling.

[NiFe] hydrogenase maturation proteins HypC, HypD, and HypE catalyze the insertion and cyanation of the iron center of [NiFe] hydrogenases by an unknown mechanism. We have determined the crystal structures of HypC, HypD, and HypE from Thermococcus kodakaraensis KOD1 at 1.8 A, 2.

Crystallization and preliminary X-ray crystallographic studies of the [NiFe] hydrogenase maturation proteins HypC and HypD.

HypC and HypD proteins are required for the insertion of the Fe atom with diatomic ligands into the large subunit of [NiFe] hydrogenases, an important step in the maturation process of this type of hydrogenase. The crystallization and preliminary crystallographic analysis of HypC and HypD from Thermococcus kodakaraensis KOD1 are reported. Crystals of HypC grew in two different forms.

Disruption of a sugar transporter gene cluster in a hyperthermophilic archaeon using a host-marker system based on antibiotic resistance.

We have developed a gene disruption system in the hyperthermophilic archaeon Thermococcus kodakaraensis using the antibiotic simvastatin and a fusion gene designed to overexpress the 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase gene (hmg(Tk)) with the glutamate dehydrogenase promoter. With this system, we disrupted the T. kodakaraensis amylopullulanase gene (apu(Tk)) or a gene cluster which includes apu(Tk) and genes encoding components of a putative sugar transporter.

Isolation and characterization of a novel poly(vinyl alcohol)-degrading bacterium, Sphingopyxis sp. PVA3.

We have isolated a poly(vinyl alcohol) (PVA)-degrading bacterium from an activated sludge sample obtained from the drainage of a dyeing factory. Enrichment cultures were performed in media containing PVA as the sole or major carbon source. After several rounds of cultivation on liquid and solid media, we were able to isolate a single colony with PVA-degrading ability (strain PVA3).

Identification of the amino acid residues essential for proteolytic activity in an archaeal signal peptide peptidase.

Signal peptide peptidases (SPPs) are enzymes involved in the initial degradation of signal peptides after they are released from the precursor proteins by signal peptidases. In contrast to the eukaryotic enzymes that are aspartate peptidases, the catalytic mechanisms of prokaryotic SPPs had not been known. In this study on the SPP from the hyperthermophilic archaeon Thermococcus kodakaraensis (SppA(Tk)), we have identified amino acid residues that are essential for the peptidase activity of the enzyme.

Biochemical properties of a putative signal peptide peptidase from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1.

We have performed the first biochemical characterization of a putative archaeal signal peptide peptidase (SppA(Tk)) from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. SppA(Tk), comprised of 334 residues, was much smaller than its counterpart from Escherichia coli (618 residues) and harbored a single predicted transmembrane domain near its N terminus. A truncated mutant protein without the N-terminal 54 amino acid residues (deltaN54SppA(Tk)) was found to be stable against autoproteolysis and was examined further.

Complete genome sequence of the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 and comparison with Pyrococcus genomes.

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

Reverse gyrase is not a prerequisite for hyperthermophilic life.

We disrupted the reverse gyrase gene from a hyperthermophilic archaeon, Thermococcus kodakaraensis KOD1. An apparent positive supercoiling activity that was observed in the host strain was not found in the disruptant strain. We found that a lack of reverse gyrase led to a retardation in growth that was more striking at higher temperatures.