This service exclusively searches for literature that cites resources. Please be aware that the total number of searchable documents is limited to those containing RRIDs and does not include all open-access literature.
A high level of transposon-mediated genome rearrangement is a common trait among microorganisms isolated from thermal environments, probably contributing to the extraordinary genomic plasticity and horizontal gene transfer (HGT) observed in these habitats. In this work, active and inactive insertion sequences (ISs) spanning the sequenced members of the genus Thermus were characterized, with special emphasis on three T. thermophilus strains: HB27, HB8, and NAR1. A large number of full ISs and fragments derived from different IS families were found, concentrating within megaplasmids present in most isolates. Potentially active ISs were identified through analysis of transposase integrity, and domestication-related transposition events of ISTth7 were identified in laboratory-adapted HB27 derivatives. Many partial copies of ISs appeared throughout the genome, which may serve as specific targets for homologous recombination contributing to genome rearrangement. Moreover, recruitment of IS1000 32 bp segments as spacers for CRISPR sequence was identified, pointing to the adaptability of these elements in the biology of these thermophiles. Further knowledge about the activity and functional diversity of ISs in this genus may contribute to the generation of engineered transposons as new genetic tools, and enrich our understanding of the outstanding plasticity shown by these thermophiles.
Thermus aquaticus Y51MC23 was isolated from a boiling spring in the Lower Geyser Basin of Yellowstone National Park. Remarkably, this T. aquaticus strain is able to grow anaerobically and produces multiple morphological forms. Y51MC23 is a Gram-negative, rod-shaped organism that grows well between 50°C and 80°C with maximum growth rate at 65°C to 70°C. Growth studies suggest that Y51MC23 primarily scavenges protein from the environment, supported by the high number of secreted and intracellular proteases and peptidases as well as transporter systems for amino acids and peptides. The genome was assembled de novo using a 350 bp fragment library (paired end sequencing) and an 8 kb long span mate pair library. A closed and finished genome was obtained consisting of a single chromosome of 2.15 Mb and four plasmids of 11, 14, 70, and 79 kb. Unlike other Thermus species, functions usually found on megaplasmids were identified on the chromosome. The Y51MC23 genome contains two full and two partial prophage as well as numerous CRISPR loci. The high identity and synteny between Y51MC23 prophage 2 and that of Thermus sp. 2.9 is interesting, given the 8,800 km separation of the two hot springs from which they were isolated. The anaerobic lifestyle of Y51MC23 is complex, with multiple morphologies present in cultures. The use of fluorescence microscopy reveals new details about these unusual morphological features, including the presence of multiple types of large and small spheres, often forming a confluent layer of spheres. Many of the spheres appear to be formed not from cell envelope or outer membrane components as previously believed, but from a remodeled peptidoglycan cell wall. These complex morphological forms may serve multiple functions in the survival of the organism, including food and nucleic acid storage as well as colony attachment and organization.
The coupling between the binding of the substrate Fru-6-P and the inhibitor phospho(enol)pyruvate (PEP) in phosphofructokinase (PFK) from the extreme thermophile Thermus thermophilus is much weaker than that seen in a PFK from Bacillus stearothermophilus. From the crystal structures of Bacillus stearothermophilus PFK (BsPFK) the residues at positions 59, 158, and 215 in BsPFK are located on the path leading from the allosteric site to the nearest active site and are part of the intricate hydrogen-bonding network connecting the two sites. Substituting the corresponding residues in Thermus thermophilus PFK (TtPFK) with the amino acids found at these positions in BsPFK allowed us to enhance the allosteric inhibition by PEP by nearly 3 kcal mol(-1) (50-fold) to a value greater than or equal to the coupling observed in BsPFK. Interestingly, each single variant N59D, A158T, and S215H produced a roughly 1 kcal mol(-1) increase in coupling free energy of inhibition. The effects of these variants were essentially additive in the three combinations of double variants N59D/A158T, N59D/S215H, and A158T/S215H as well as in the triple variant N59D/A158T/S215H. Consequently, while the hydrogen-bonding network identified is likely involved in the inhibitory allosteric communication, a model requiring a linked chain of interactions connecting the sites is not supported by these data. Despite the fact that the allosteric activator of the bacterial PFK, MgADP, binds at the same allosteric site, the substitutions at positions 59, 158, and 215 do not have an equally dramatic effect on the binding affinity and the allosteric activation by MgADP. The effect of the S215H and N59D/A158T/S215H substitutions on the activation by MgADP could not be determined because of a dramatic drop in MgADP binding affinity that resulted from the S215H substitution. The single variants N59D and A158T supported binding but showed little change in the free energy of activation by MgADP compared to the wild type TtPFK. These results support previous suggestions that heterotropic inhibition and activation occur by different pathways prokaryotic PFK.
Genome sequencing of the yellow-pigmented, thermophilic bacterium Thermus sp. NMX2.A1 resulted in a 2.29 Mb draft genome that encodes for 2312 proteins. The genetic relationship between various strains from the genus Thermus was assessed based on phylogenomic analyses using a concatenated set of conserved proteins. The resulting phylogenetic tree illustrated that Thermus sp. NMX2 A.1 clusters together with Thermus scotoductus SA-01, despite being isolated from vastly different geographical locations. The close evolutionary relationship and metabolic parallels between the two strains has previously been recognized; however, neither strain's genome data were available at that point in time. Genomic comparison of the Thermus sp. NMX2.A1 and T. scotoductus SA-01, as well as other closely related Thermus strains, revealed a high degree of synteny at both the genomic and proteomic level, with processes such as denitrification and natural cell competence appearing to be conserved. However, despite this high level of similarity, analysis revealed a complete, putative Calvin-Benson-Bassham (CBB) cycle in NMX2.A1 that is absent in SA-01. Analysis of horizontally transferred gene islands provide evidence that NMX2 selected these genes due to pressure from its HCO3 (-) rich environment, which is in stark contrast to that of the deep subsurface isolated SA-01.
The Thermus thermophilus succinate:quinone reductase (SQR), serving as the respiratory complex II, has been homologously produced under the control of a constitutive promoter and subsequently purified. The detailed biochemical characterization of the resulting wild type (wt-rcII) and His-tagged (rcII-His(8)-SdhB and rcII-SdhB-His(6)) complex II variants showed the same properties as the native enzyme with respect to the subunit composition, redox cofactor content and sensitivity to the inhibitors malonate, oxaloacetate, 3-nitropropionic acid and nonyl-4-hydroxyquinoline-N-oxide (NQNO). The position of the His-tag determined whether the enzyme retained its native trimeric conformation or whether it was present in a monomeric form. Only the trimer exhibited positive cooperativity at high temperatures. The EPR signal of the [2Fe-2S] cluster was sensitive to the presence of substrate and showed an increased rhombicity in the presence of succinate in the native and in all recombinant forms of the enzyme. The detailed analysis of the shape of this signal as a function of pH, substrate concentration and in the presence of various inhibitors and quinones is presented, leading to a model for the molecular mechanism that underlies the influence of succinate on the rhombicity of the EPR signal of the proximal iron-sulfur cluster.
A DNA sequencing-based strategy was applied to study the microbiology of Continental-type cheeses with a pink discoloration defect. The basis for this phenomenon has remained elusive, despite decades of research. The bacterial composition of cheese containing the defect was compared to that of control cheese using 16S rRNA gene and shotgun metagenomic sequencing as well as quantitative PCR (qPCR). Throughout, it was apparent that Thermus, a carotenoid-producing genus, was present at higher levels in defect-associated cheeses than in control cheeses. Prompted by this finding and data confirming the pink discoloration to be associated with the presence of a carotenoid, a culture-based approach was employed, and Thermus thermophilus was successfully cultured from defect-containing cheeses. The link between Thermus and the pinking phenomenon was then established through the cheese defect equivalent of Koch's postulates when the defect was recreated by the reintroduction of a T. thermophilus isolate to a test cheese during the manufacturing process. IMPORTANCE Pink discoloration in cheese is a defect affecting many cheeses throughout the world, leading to significant financial loss for the dairy industry. Despite decades of research, the cause of this defect has remained elusive. The advent of high-throughput, next-generation sequencing has revolutionized the field of food microbiology and, with respect to this study, provided a means of testing a possible microbial basis for this defect. In this study, a combined 16S rRNA, whole-genome sequencing, and quantitative PCR approach was taken. This resulted in the identification of Thermus, a carotenoid-producing thermophile, in defect-associated cheeses and the recreation of the problem in cheeses to which Thermus was added. This finding has the potential to lead to new strategies to eliminate this defect, and our method represents an approach that can be employed to investigate the role of microbes in other food defects of unknown origin.
The extremely radioresistant bacteria of the genus Deinococcus and the extremely thermophilic bacteria of the genus Thermus belong to a common taxonomic group. Considering the distinct living environments of Deinococcus and Thermus, different genes would have been acquired through horizontal gene transfer after their divergence from a common ancestor. Their guanine-cytosine (GC) contents are similar; however, we hypothesized that their genomic signatures would be different. Our findings indicated that the genomes of Deinococcus radiodurans and Thermus thermophilus have different tetranucleotide frequencies. This analysis showed that the genome signature of D. radiodurans is most similar to that of Pseudomonas aeruginosa, whereas the genome signature of T. thermophilus is most similar to that of Thermanaerovibrio acidaminovorans. This difference in genome signatures may be related to the different evolutionary backgrounds of the 2 genera after their divergence from a common ancestor.
Transcription elongation factors from the NusG family are ubiquitous from bacteria to humans and play diverse roles in the regulation of gene expression. These proteins consist of at least two domains. The N-terminal domains directly bind to the largest, β' in bacteria, subunit of RNA polymerase (RNAP), whereas the C-terminal domains interact with other cellular components and serve as platforms for the assembly of large nucleoprotein complexes. Escherichia coli NusG and its paralog RfaH modify RNAP into a fast, pause-resistant state but the detailed molecular mechanism of this modification remains unclear since no high-resolution structural data are available for the E. coli system. We wanted to investigate whether Thermus thermophilus (Tth) NusG can be used as a model for structural studies of this family of regulators. Here, we show that Tth NusG slows down rather than facilitates transcript elongation by its cognate RNAP. On the other hand, similarly to the E. coli regulators, Tth NusG apparently binds near the upstream end of the transcription bubble, competes with σ(A), and favors forward translocation by RNAP. Our data suggest that the mechanism of NusG recruitment to RNAP is universally conserved even though the regulatory outcomes among its homologs may appear distinct.
Eukaryotic Argonaute proteins mediate RNA-guided RNA interference, allowing both regulation of host gene expression and defense against invading mobile genetic elements. Recently, it has become evident that prokaryotic Argonaute homologs mediate DNA-guided DNA interference, and play a role in host defense. Argonaute of the bacterium Thermus thermophilus (TtAgo) targets invading plasmid DNA during and after transformation. Using small interfering DNA guides, TtAgo can cleave single and double stranded DNAs. Although TtAgo additionally has been demonstrated to cleave RNA targets complementary to its DNA guide in vitro, RNA targeting by TtAgo has not been demonstrated in vivo.
Phosphoribosyltransferases are the tools that allow the synthesis of nucleotide analogues using multi-enzymatic cascades. The recombinant adenine phosphoribosyltransferase (TthAPRT) and hypoxanthine phosphoribosyltransferase (TthHPRT) from Thermus thermophilus HB27 were expressed in E.coli strains and purified by chromatographic methods with yields of 10-13 mg per liter of culture. The activity dependence of TthAPRT and TthHPRT on different factors was investigated along with the substrate specificity towards different heterocyclic bases. The kinetic parameters for TthHPRT with natural substrates were determined. Two nucleotides were synthesized: 9-(β-D-ribofuranosyl)-2-chloroadenine 5'-monophosphate (2-Сl-AMP) using TthAPRT and 1-(β-D-ribofuranosyl)pyrazolo[3,4-d]pyrimidine-4-one 5'-monophosphate (Allop-MP) using TthНPRT.
Tiamulin is a semisynthetic pleuromutilin antibiotic that binds to the 50S ribosomal subunit A site and whose (((2-diethylamino)ethyl)thio)-acetic acid tail extends into the P site to interfere with peptide bond formation. We have isolated spontaneous tiamulin-resistant mutants of the thermophilic bacterium Thermus thermophilus, containing either single amino acid substitutions in ribosomal protein uL3 or single base substitutions in the peptidyltransferase active site of 23S rRNA. These mutations are consistent with those found in other organisms and are in close proximity to the crystallographically determined tiamulin binding site. We also conducted a cross-resistance analysis of nine other single-base substitutions in or near the peptidyltransferase active site, previously selected for resistance to structurally unrelated antibiotics. While some of the base substitutions in 23S rRNA are positioned to directly affect tiamulin-ribosome contacts, others are some distance from the tiamulin binding site, indicating an indirect mechanism of resistance. Similarly, amino acid substitutions in uL3 are predicted to act indirectly by destabilizing rRNA conformation in the active site. We interpret these observations in light of the available ribosome X-ray crystal structures. These results provide a more comprehensive profile of tiamulin resistance caused by mutations in the bacterial ribosome.
Thermus thermophilus trigger factor (TtTF) is a zinc-dependent molecular chaperone whose folding-arrest activity is regulated by Zn2+. However, little is known about the mechanism of zinc-dependent regulation of the TtTF activity. Here we exploit in vitro biophysical experiments to investigate zinc-binding, the oligomeric state, the secondary structure, and the thermal stability of TtTF in the absence and presence of Zn2+. The data show that full-length TtTF binds Zn2+, but the isolated domains and tandem domains of TtTF do not bind to Zn2+. Furthermore, circular dichroism (CD) and nuclear magnetic resonance (NMR) spectra suggested that Zn2+-binding induces the partial structural changes of TtTF, and size exclusion chromatography-multi-angle light scattering (SEC-MALS) showed that Zn2+ promotes TtTF oligomerization. Given the previous work showing that the activity regulation of E. coli trigger factor is accompanied by oligomerization, the data suggest that TtTF exploits zinc ions to induce the structural change coupled with the oligomerization to assemble the client-binding site, thereby effectively preventing proteins from misfolding in the thermal environment.
Isoprenoid precursor biosynthesis occurs through the mevalonate or the methylerythritol phosphate (MEP) pathway, used i.e., by humans and by many human pathogens, respectively. In the MEP pathway, 2-C-methyl-D-erythritol-2,4-cyclo-diphosphate (MEcPP) is converted to (E)-1-hydroxy-2-methyl-but-2-enyl-4-diphosphate (HMBPP) by the iron-sulfur cluster enzyme HMBPP synthase (GcpE). The presented X-ray structure of the GcpE-MEcPP complex from Thermus thermophilus at 1.55Å resolution provides valuable information about the catalytic mechanism and for rational inhibitor design. MEcPP binding inside the TIM-barrel funnel induces a 60° rotation of the [4Fe-4S] cluster containing domain onto the TIM-barrel entrance. The apical iron of the [4Fe-4S] cluster ligates with the C3 oxygen atom of MEcPP.
This study describes the structure of DNA polymerase I from Thermus phage G20c, termed PolI_G20c. This is the first structure of a DNA polymerase originating from a group of related thermophilic bacteriophages infecting Thermus thermophilus, including phages G20c, TSP4, P74-26, P23-45 and phiFA and the novel phage Tth15-6. Sequence and structural analysis of PolI_G20c revealed a 3'-5' exonuclease domain and a DNA polymerase domain, and activity screening confirmed that both domains were functional. No functional 5'-3' exonuclease domain was present. Structural analysis also revealed a novel specific structure motif, here termed SβαR, that was not previously identified in any polymerase belonging to the DNA polymerases I (or the DNA polymerase A family). The SβαR motif did not show any homology to the sequences or structures of known DNA polymerases. The exception was the sequence conservation of the residues in this motif in putative DNA polymerases encoded in the genomes of a group of thermophilic phages related to Thermus phage G20c. The structure of PolI_G20c was determined with the aid of another structure that was determined in parallel and was used as a model for molecular replacement. This other structure was of a 3'-5' exonuclease termed ExnV1. The cloned and expressed gene encoding ExnV1 was isolated from a thermophilic virus metagenome that was collected from several hot springs in Iceland. The structure of ExnV1, which contains the novel SβαR motif, was first determined to 2.19 Å resolution. With these data at hand, the structure of PolI_G20c was determined to 2.97 Å resolution. The structures of PolI_G20c and ExnV1 are most similar to those of the Klenow fragment of DNA polymerase I (PDB entry 2kzz) from Escherichia coli, DNA polymerase I from Geobacillus stearothermophilus (PDB entry 1knc) and Taq polymerase (PDB entry 1bgx) from Thermus aquaticus.
Despite the fact that DNA polymerases have been investigated for many years and are commonly used as tools in a number of molecular biology assays, many details of the kinetic mechanism they use to catalyze DNA synthesis remain unclear. Structural and kinetic studies have characterized a rapid, pre-catalytic open-to-close conformational change of the Finger domain during nucleotide binding for many DNA polymerases including Thermus aquaticus DNA polymerase I (Taq Pol), a thermostable enzyme commonly used for DNA amplification in PCR. However, little has been performed to characterize the motions of other structural domains of Taq Pol or any other DNA polymerase during catalysis. Here, we used stopped-flow Förster resonance energy transfer to investigate the conformational dynamics of all five structural domains of the full-length Taq Pol relative to the DNA substrate during nucleotide binding and incorporation. Our study provides evidence for a rapid conformational change step induced by dNTP binding and a subsequent global conformational transition involving all domains of Taq Pol during catalysis. Additionally, our study shows that the rate of the global transition was greatly increased with the truncated form of Taq Pol lacking the N-terminal domain. Finally, we utilized a mutant of Taq Pol containing a de novo disulfide bond to demonstrate that limiting protein conformational flexibility greatly reduced the polymerization activity of Taq Pol.
Little is known about chromosome segregation in polyploid prokaryotes. In this study, whether stringent or variable chromosome segregation occurs in polyploid thermophilic bacterium Thermus thermophilus was analyzed. A stable heterozygous strain (HL01) containing two antibiotic resistance markers at one gene locus was generated. The inheritance of the two alleles in the progeny of the heterozygous strain was then followed. During incubation without selection pressure, the fraction of heterozygous cells decreased and that of homozygous cells increased, while the relative abundance of each allele in the whole population remained constant, suggesting chromosome segregation had experienced random event. Consistently, in comparison with Bacillus subtilis in which the sister chromosomes were segregated equally, the ratios of DNA content in two daughter cells of T. thermophilus had a broader distribution and a larger standard deviation, indicating that the DNA content in the two daughter cells was not always identical. Further, the protein homologs (i.e., ParA and MreB) which have been suggested to be involved in bacterial chromosome partitioning did not actively participate in the chromosome segregation in T. thermophilus Therefore, it seems that protein-based chromosome segregation machineries are less critical for the polyploid T. thermophilus, and chromosome segregation in this bacterium are not stringently controlled but tend to be variable, and random segregation can occur.
Secretins form multimeric channels across the outer membrane of Gram-negative bacteria that mediate the import or export of substrates and/or extrusion of type IV pili. The secretin complex of Thermus thermophilus is an oligomer of the 757-residue PilQ protein, essential for DNA uptake and pilus extrusion. Here, we present the cryo-EM structure of this bifunctional complex at a resolution of ~7 Å using a new reconstruction protocol. Thirteen protomers form a large periplasmic domain of six stacked rings and a secretin domain in the outer membrane. A homology model of the PilQ protein was fitted into the cryo-EM map. A crown-like structure outside the outer membrane capping the secretin was found not to be part of PilQ. Mutations in the secretin domain disrupted the crown and abolished DNA uptake, suggesting a central role of the crown in natural transformation.
Thermophilic environments represent an interesting niche. Among thermophiles, the genus Thermus is among the most studied genera. In this study, we have sequenced the genome of Thermus parvatiensis strain RL, a thermophile isolated from Himalayan hot water springs (temperature >96°C) using PacBio RSII SMRT technique. The small genome (2.01 Mbp) comprises a chromosome (1.87 Mbp) and a plasmid (143 Kbp), designated in this study as pTP143. Annotation revealed a high number of repair genes, a squeezed genome but containing highly plastic plasmid with transposases, integrases, mobile elements and hypothetical proteins (44%). We performed a comparative genomic study of the group Thermus with an aim of analysing the phylogenetic relatedness as well as niche specific attributes prevalent among the group. We compared the reference genome RL with 16 Thermus genomes to assess their phylogenetic relationships based on 16S rRNA gene sequences, average nucleotide identity (ANI), conserved marker genes (31 and 400), pan genome and tetranucleotide frequency. The core genome of the analyzed genomes contained 1,177 core genes and many singleton genes were detected in individual genomes, reflecting a conserved core but adaptive pan repertoire. We demonstrated the presence of metagenomic islands (chromosome:5, plasmid:5) by recruiting raw metagenomic data (from the same niche) against the genomic replicons of T. parvatiensis. We also dissected the CRISPR loci wide all genomes and found widespread presence of this system across Thermus genomes. Additionally, we performed a comparative analysis of competence loci wide Thermus genomes and found evidence for recent horizontal acquisition of the locus and continued dispersal among members reflecting that natural competence is a beneficial survival trait among Thermus members and its acquisition depicts unending evolution in order to accomplish optimal fitness.
In many eukaryotes, Argonaute proteins, guided by short RNA sequences, defend cells against transposons and viruses. In the eubacterium Thermus thermophilus, the DNA-guided Argonaute TtAgo defends against transformation by DNA plasmids. Here, we report that TtAgo also participates in DNA replication. In vivo, TtAgo binds 15- to 18-nt DNA guides derived from the chromosomal region where replication terminates and associates with proteins known to act in DNA replication. When gyrase, the sole T. thermophilus type II topoisomerase, is inhibited, TtAgo allows the bacterium to finish replicating its circular genome. In contrast, loss of gyrase and TtAgo activity slows growth and produces long sausage-like filaments in which the individual bacteria are linked by DNA. Finally, wild-type T. thermophilus outcompetes an otherwise isogenic strain lacking TtAgo. We propose that the primary role of TtAgo is to help T. thermophilus disentangle the catenated circular chromosomes generated by DNA replication.
The ring-shaped hexameric DnaB helicase unwinds duplex DNA at the replication fork of eubacteria. We have solved the crystal structure of the full-length Thermus aquaticus DnaB monomer, or possibly dimer, at 2.9 A resolution. DnaB is a highly flexible two domain protein. The C-terminal domain exhibits a RecA-like core fold and contains all the conserved sequence motifs that are characteristic of the DnaB helicase family. The N-terminal domain contains an additional helical hairpin that makes it larger than previously appreciated. Several DnaB mutations that modulate its interaction with primase are found in this hairpin. The similarity in the fold of the DnaB N-terminal domain with that of the C-terminal helicase-binding domain (HBD) of the DnaG primase also includes this hairpin. Comparison of hexameric homology models of DnaB with the structure of the papillomavirus E1 helicase suggests the two helicases may function through different mechanisms despite their sharing a common ancestor.
Welcome to the FDI Lab - SciCrunch.org Resources search. From here you can search through a compilation of resources used by FDI Lab - SciCrunch.org and see how data is organized within our community.
You are currently on the Community Resources tab looking through categories and sources that FDI Lab - SciCrunch.org has compiled. You can navigate through those categories from here or change to a different tab to execute your search through. Each tab gives a different perspective on data.
If you have an account on FDI Lab - SciCrunch.org then you can log in from here to get additional features in FDI Lab - SciCrunch.org such as Collections, Saved Searches, and managing Resources.
Here is the search term that is being executed, you can type in anything you want to search for. Some tips to help searching:
You can save any searches you perform for quick access to later from here.
We recognized your search term and included synonyms and inferred terms along side your term to help get the data you are looking for.
If you are logged into FDI Lab - SciCrunch.org you can add data records to your collections to create custom spreadsheets across multiple sources of data.
Here are the facets that you can filter your papers by.
From here we'll present any options for the literature, such as exporting your current results.
If you have any further questions please check out our FAQs Page to ask questions and see our tutorials. Click this button to view this tutorial again.
Year:
Count: