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PubMed1988

Characterization of 15 selected coccal bacteria isolated from Antarctic rock and soil samples from the McMurdo-Dry Valleys (South-Victoria Land).

Approximately 1500 cultures of microorganisms were isolated from rocks and soils of the Ross Desert (McMurdo-Dry Valleys). From these, 15 coccoid strains were chosen for more detailed investigation. They were characterized by morphological, physiological and chemotaxonomical properties. All isolates were Gram-positive, catalase-positive and nonmotile. Six strains showed red pigmentation and could be identified as members of the genera Micrococcus (M. roseus, M. agilis) or Deinococcus. In spite of their coccoid morphology, the remaining nine strains had to be associated with coryneform bacteria (Arthrobacter, Brevibacterium), because of their cell wall composition and G+C ratios. Most of the strains were psychrotrophic, but one strain was even obligately psychrophilic, with a temperature maximum below 20 degrees C. Red cocci had in vitro pH optima above 9.0 although they generally originated from acid samples. Most isolates showed a preference for sugar alcohols and organic acids, compounds which are commonly known to be released by lichens, molds and algae, the other components of the cryptoendolithic ecosystem. These properties indicate that our strains are autochthonous members of the natural Antarctic microbial population.

Antarctic Regions↗
PubMed1991

DNA-strand breaks limit survival in extreme dryness.

The inactivation of the anhydrobiotic organisms Bacillus subtilis (spores) and Deinococcus radiodurans during long-term exposure (up to several weeks) to extreme dryness (especially vacuum) is correlated with an increase in the number of DNA-strand breaks and other DNA lesions. Survival finally depends on the repair of DNA damages. Exposure of anhydrobiotic organisms to extreme dryness (e.g. on Mars or in space) for geological times will lead to so extended DNA lesions that recovery is extremely unlikely.

Bacillus subtilis↗
PubMed1999

DNA polymerase C of the thermophilic bacterium Thermus aquaticus: classification and phylogenetic analysis of the family C DNA polymerases.

Bacterial family C DNA polymerases (DNA pol IIIs), the major chromosomal replicative enzymes, have been provisionally classified based on primary sequences and domain structures into three classes: class I (Escherichia coli DNA pol C-type), class II (Bacillus subtilis DNA pol C-type), and class III (cyanobacterial DNA pol C-type), respectively. We have sequenced the structural gene encoding the DNA pol C catalytic subunit of the thermophilic bacterium Thermus aquaticus. This gene, designated the Taq DNA pol C gene, contains a 3660-bp open reading frame which specifies a polypeptide of molecular weight of 137,388 daltons. Comparative sequence analyses revealed that Taq DNA pol C is a class I family C DNA polymerase. The Taq DNA pol C is most closely related to the Deinococcus radiodurans DNA pol C. Although a phylogenetic tree based on the class I family C DNA pols is still in the provisional stage, some important conclusion can be drawn. First, the high-G+C and the low-G+C Gram-positive bacteria are not monophyletic. Second, the low-G+C Gram-positive bacteria contain multigenes of family C DNA pols (classes I and II). Third, the cyanobacterial family C DNA pol, classified as class III because it is encoded by a split gene, forms a group with the high-G+C Gram-positive bacteria.

Amino Acid Sequence↗
PubMed2000

A new appraisal of the prokaryotic origin of eukaryotic phytochromes.

The evolutionary origin of the phytochromes of eukaryotes is controversial. Three cyanobacterial proteins have been described as "phytochrome-like" and have been suggested to be potential ancestors of these essential photoreceptors: Cph1 from Synechocystis PCC 6803, showing homology to phytochromes along its entire length and known to attach a chromophore; and PlpA from Synechocystis PCC 6803 and RcaE from Fremyella diplosiphon, both showing homology to phytochromes most strongly only in the C-terminal region and not known to bind a chromophore. We have reexamined the evolution of the photoreceptors using for PCR amplification a highly conserved region encoding the chromophore-binding domain in both Cph1 and phytochromes of plants and have identified genes for phytochrome-like proteins (PLP) in 11 very diverse cyanobacteria. The predicted gene products contain either a Cys, Arg, Ile, or Leu residue at the putative chromophore binding site. In 10 of the strains examined only a single gene was found, but in Calothrix PCC 7601 two genes (cphA and cphB) were identified. Phylogenetic analysis revealed that genes encoding PLP are homologues that share a common ancestor with the phytochromes of eukaryotes and diverged before the latter. In contrast, the putative sensory/regulatory proteins, including PlpA and RcaE, that lack a part of the chromophore lyase domain essential for chromophore attachment on the apophytochrome, are only distantly related to phytochromes. The Ppr protein of the anoxygenic photosynthetic bacterium Rhodospirillum centenum and the bacterial phytochrome-like proteins (BphP) of Deinococcus radiodurans and Pseudomonas aeruginosa fall within the cluster of cyanobacterial phytochromes.

Amino Acid Sequence↗
PubMed2000

Horizontal transfer of archaeal genes into the deinococcaceae: detection by molecular and computer-based approaches.

Members of the Deinococcaceae (e.g., Thermus, Meiothermus, Deinococcus) contain A/V-ATPases typically found in Archaea or Eukaryotes which were probably acquired by horizontal gene transfer. Two methods were used to quantify the extent to which archaeal or eukaryotic genes have been acquired by this lineage. Screening of a Meiothermus ruber library with probes made against Thermoplasma acidophilum DNA yielded a number of clones which hybridized more strongly than background. One of these contained the prolyl tRNA synthetase (RS) gene. Phylogenetic analysis shows the M. ruber and D. radiodurans prolyl RS to be more closely related to archaeal and eukaryal forms of this gene than to the typical bacterial type. Using a bioinformatics approach, putative open reading frames (ORFs) from the prerelease version of the D. radiodurans genome were screened for genes more closely related to archaeal or eukaryotic genes. Putative ORFs were searched against representative genomes from each of the three domains using automated BLAST. ORFs showing the highest matches against archaeal and eukaryotic genes were collected and ranked. Among the top-ranked hits were the A/V-ATPase catalytic and noncatalytic subunits and the prolyl RS genes. Using phylogenetic methods, ORFs were analyzed and trees assessed for evidence of horizontal gene transfer. Of the 45 genes examined, 20 showed topologies in which D. radiodurans homologues clearly group with eukaryotic or archaeal homologues, and 17 additional trees were found to show probable evidence of horizontal gene transfer. Compared to the total number of ORFs in the genome, those that can be identified as having been acquired from Archaea or Eukaryotes are relatively few (approximately 1%), suggesting that interdomain transfer is rare.

Archaea↗
PubMed2004

The antioxidative effect of the bacteria Dienococcus radiophilus against LDL lipid peroxidation.

BACKGROUND: Lipid peroxidation is an important process in the development of atherosclerosis. Thus agents with antioxidant properties may play an important role in the inhibition of atherosclerosis. OBJECTIVES: In this study we aimed to show that the lipid extract of the bacteria Deinococcus radiophilus (leDR) has antioxidant properties against LDL oxidation. RESULTS: This antioxidant effect was shown in both transition metal ion and free radical generating systems. We also showed that leDR can protect LDL from UV light-induced oxidative damage. The antioxidative capacity of leDR is partly due to copper ion chelation. CONCLUSION: We conclude that some specific bacteria constituent has the ability to inhibit LDL oxidation and, thus, to attenuate atherogenesis.

Atherosclerosis↗
PubMed2001

DNA protection mechanisms are not involved in the radioresistance of the hyperthermophilic archaea Pyrococcus abyssi and P. furiosus.

Hyperthermophilic archaea of the genus Pyrococcus are resistant to gamma radiation, suggesting that efficient mechanisms for DNA repair exist in these organisms. To determine whether protective mechanisms might also be implicated in this radioresistance, we have estimated the linear density of DNA double-stranded breaks caused by gamma irradiation in the genomic DNA of two Pyrococcus species, using Escherichia coli and the radioresistant bacterium Deinococcus radiodurans as controls. The linear density of double-stranded breaks was essentially the same in all four microorganisms when irradiation was carried under similar anaerobic conditions, indicating that no specific DNA protection mechanisms exist in Pyrococcus species. Using one- and two-dimensional gel electrophoresis we compared the protein patterns from Pyrococcus abyssi and P. furiosus cells that had or had not been exposed to gamma rays. We did not detect any significant protein induction following DNA damage in either species.

DNA Damage↗
PubMed2005

Isolation and characterization of Thermus bacteriophages.

One-hundred-fifteen bacteriophage strains were isolated from alkaline hot springs in Iceland, New Zealand, Russia (Kamchatka), and the U.S.A. The phages belonged to the Myoviridae, Siphoviridae, Tectiviridae, and Inoviridae families. Over 50% of isolates were isometric or filamentous. One type of siphovirus had giant tails of over 800 nm in length. Phages were further characterized by host range, genome size, DNA restriction endonuclease digestion patterns, and temperature and pH sensitivity. Myoviruses and tectiviruses had a worldwide distribution. Most phages were narrowly host-specific and all were highly resistant against heating and alkaline and acidic pH. This is the first time that tectiviruses and filamentous phages are reported for bacteria of the Thermus-Deinococcus phylum. The presence of tectiviruses, inoviruses, and myoviruses is attributed to acquisition from ancestral gamma-proteobacteria by horizontal gene transfer.

Alkalies↗
PubMed2002

Citrate synthase from Thermus aquaticus: a thermostable bacterial enzyme with a five-membered inter-subunit ionic network.

A bacterial thermostable citrate synthase has been analyzed to investigate the structural basis of its thermostability, and to compare such features with those previously identified in archaeal citrate synthases. The gene encoding the citrate synthase from Thermus aquaticus was identified from a gene library by screening with a PCR fragment amplified from genomic DNA using a primer based on the determined N-terminal amino acid sequence and a citrate synthase consensus primer. Apart from high sequence similarities with citrate synthase sequences within the Thermus/ Deinococcus group, the analyzed enzyme has highest similarities with the enzyme from the hyperthermophilic Archaeon Pyrococcus furiosus. The recombinant enzyme is a dimer with high specific activity. Compared to its thermoactivity (T(opt)at 80 degrees C), the thermal stability of the enzyme is high, as judged from its T(m) (101 degrees C), and from irreversible thermal inactivation assays. Molecular modeling of the structure revealed an inter-subunit ion-pair network, comparable in size to the network found in the citrate synthase from P. furiosus; these networks are discussed in relation to the high thermal stability of these bacterial and archaeal enzymes.

Amino Acid Sequence↗
PubMed2001

The branching order and phylogenetic placement of species from completed bacterial genomes, based on conserved indels found in various proteins.

The presence of shared conserved inserts and deletions (indels or signature sequences) in proteins provides a powerful means for understanding the evolutionary relationships among the Bacteria. Using such indels, all of the main groups within the Bacteria can be defined in clear molecular terms and it has become possible to deduce that they branched from a common ancestor in the following order: Low G + C gram-positive --> High G+C gram-positive --> Deinococcus Thermus --> Cyanobacteria --> Spirochetes --> Aquifex-Chlamydia-Cytophaga --> Proteobacteria-1 (epsilon, delta) --> Proteobacteria-2 (alpha) --> Proteobacteria-3 (beta) --> Proteobacteria -4 (gamma). The usefulness of this approach for understanding bacterial phylogeny was examined here using sequence data from various completed bacterial genomes. By using 12 indels in highly conserved and widely represented proteins, the species from all 41 completed bacterial genomes were assigned to different groups; and the observed distribution of these indels in different species was then compared with that predicted by the signature sequence model. The presence or absence of these indels in various proteins in different bacteria followed the pattern exactly as predicted: and, in more than 450 observations, no exceptions or contradictions in the placement of indels were observed. These results provide strong evidence that lateral gene transfer events have not affected the genes containing these indels to any significant extent. The phylogenetic placement of bacteria into different groups based on signature sequences also showed an excellent correlation with the 16 S rRNA with 39 of the 41 species assigned to the same group by both methods. These results strongly vindicate the usefulness of the signature sequence approach to understanding phylogeny within the Bacteria and show that it provides a reliable and internally consistent means for the placement of bacterial species into different groups and for determining the relative branching order of the groups.

Amino Acid Sequence↗
PubMed2006

Microbial community of a hydrothermal mud vent underneath the deep-sea anoxic brine lake Urania (eastern Mediterranean).

The composition of a metabolically active prokaryotic community thriving in hydrothermal mud fluids of the deep-sea hypersaline anoxic Western Urania Basin was characterized using rRNA-based phylogenetic analysis of a clone library. The physiologically active prokaryotic assemblage in this extreme environment showed a great genetic diversity. Most members of the microbial community appeared to be affiliated to yet uncultured organisms from similar ecosystems, i.e., deep-sea hypersaline basins and hydrothermal vents. The bacterial clone library was dominated by phylotypes affiliated with the epsilon-Proteobacteria subdivision recognized as an ecologically significant group of bacteria inhabiting deep-sea hydrothermal environments. Almost 18% of all bacterial clones were related to delta-Proteobacteria, suggesting that sulfate reduction is one of the dominant metabolic processes occurring in warm mud fluids. The remaining bacterial phylotypes were related to alpha- and beta-Proteobacteria, Actinobacteria, Bacteroides, Deinococcus-Thermus, KB1 and OP-11 candidate divisions. Moreover, a novel monophyletic clade, deeply branched with unaffiliated 16S rDNA clones was also retrieved from deep-sea sediments and halocline of Urania Basin. Archaeal diversity was much lower and detected phylotypes included organisms affiliated exclusively with the Euryarchaeota. More than 96% of the archaeal clones belonged to the MSBL-1 candidate order recently found in hypersaline anoxic environments, such as endoevaporitic microbial mats, Mediterranean deep-sea mud volcanoes and anoxic basins. Two phylotypes, represented by single clones were related to uncultured groups DHVE-1 and ANME-1. Thus, the hydrothermal mud of hypersaline Urania Basin seems to contain new microbial diversity. The prokaryotic community was significantly different from that occurring in the upper layers of the Urania Basin since 60% of all bacterial and 40% of all archaeal phylotypes were obtained only from mud fluids. The uniqueness of the composition of the active prokaryotic community could be explained by the complex environmental conditions at the site. The interaction of oxygenated warm mud fluids with the cold hypersaline brine of the Urania Basin seems to simultaneously select for various metabolic processes, such as aerobic and anaerobic heterotrophy, sulfide- and methane-dependent chemotrophy along with anaerobic oxidation of methane, sulfate- and metal-reduction.

Cloning, Molecular↗
PubMedDate not supplied

Lack of ultraviolet mutagenesis in radiation-resistant bacteria.

Ultraviolet (UV) radiation did not induce rifampicin-resistant mutants in populations of the taxonomically-related radiation-resistant bacteria Deinococcus radiodurans, D. radiopugnans, D. radiophilus and D. proteolyticus, although such mutants arose spontaneously at a low frequency and at a high frequency after treatment of cultures with N-nitroso compounds. The radiation-resistant bacteria Arthrobacter radiotolerans and P-30-A were also UV-immutable whereas the more radiation-sensitive Pseudomonas radiora was UV-mutable. We conclude that the radiation-resistant bacteria repair UV-induced DNA damage accurately and lack an error-prone pathway for the repair of such damage.

Bacteria↗
PubMed1995

Scanning probe microscopy in microbiology.

Scanning probe microscopy (SPM) is emerging as an important alternative to electron microscopy as a technique for analyzing submicron details on biological surfaces. Microbiological specimens such as viruses, bacteriophages, and ordered bacterial surface layers and membranes have played an important role in the development of scanning tunnelling microscopy (STM) and atomic force microscopy (AFM) in cellular and molecular biology. Early STM studies involving metal-coated bacteriophage T4 polyheads, Methanospirillum hungatei, and Deinococcus radiodurans HPI layer clearly demonstrated that resolution was comparable to TEM on similarly prepared specimens and only limited by metal graininess. However, except for thin films or layers, novel biological information has been difficult to obtain since imaging native surfaces of such biomaterials as proteins or nucleic acids by STM proved to be unreliable. With the development of atomic force microscopes, which allow imaging of similar native structures, SPM applications have widened to include straightforward surface structure analysis, analysis of surface elastic and inelastic properties, bonding force measurements between molecules, and micro-manipulations of such individual molecules as DNA. AFM images have progressed from relatively crude representations of specimen topography to nanometer scale representations of native hydrated surfaces. It appears from the study of microbiological specimens that direct visualization of dynamic molecular events or processes may soon become a reality.

Bacteria↗
PubMed2003

The Ro autoantigen binds misfolded U2 small nuclear RNAs and assists mammalian cell survival after UV irradiation.

The Ro 60 kDa autoantigen, an RNA binding protein, is a major target of the immune response in patients with systemic lupus erythematosus. As mice lacking Ro develop a lupus-like syndrome, Ro may be important for preventing autoimmunity. However, the cellular function of Ro, which binds small cytoplasmic RNAs of unknown function called Y RNAs, has been enigmatic. Ro has been proposed to function in 5S rRNA quality control based on experiments in Xenopus laevis oocytes, and a Ro ortholog enhances survival of the eubacterium Deinococcus radiodurans after ultraviolet irradiation. To test the general importance of these two observations for Ro function, we investigated the role of Ro in mammalian cells. We report that, in mouse embryonic stem (ES) cells, Ro binds variant spliceosomal U2 snRNAs. Expression of mouse U2 snRNAs in Xenopus oocytes reveals that binding occurs in nuclei and appears to involve recognition of misfolded RNA. Moreover, mouse ES cells lacking Ro exhibit decreased survival after ultraviolet irradiation. In irradiated cells, both Ro and a Y RNA accumulate in nuclei. We propose that Ro plays a general role in small RNA quality control and that this function is important for cell survival after ultraviolet irradiation.

Animals↗
PubMed2005

Molecular phylogenetic diversity of bacteria associated with the leachate of a closed municipal solid waste landfill.

A 16S rDNA-based molecular study was performed to determine the nature of the bacterial constituents of the leachate from a closed municipal solid waste landfill. Total community DNA was extracted and bacterial 16S rRNA genes were subsequently amplified and cloned. Recombinant rDNA clones in the library were randomly selected, and they were sequenced for a single run and then grouped. A total of 76 sequence types representing 138 randomly selected nonchimeric clones were identified. Full-length sequencing and phylogenetic analysis of the sequence types revealed that more than 90% of the screened clones were affiliated with low-G+C gram-positive bacteria (38.4%), Proteobacteria (35.5%), the Cytophaga Flexibacter Bacteroides group (11.6%), and Spirochaetes (5.1%). Minor portions were affiliated with Verrucomicrobia (2.9%), candidate division OP11 (2.2%), and the green nonsulfur bacteria, Cyanobacteria and the Deinococcus Thermus group (each <1.0%). Although some rDNA sequences clustered with genera or taxa that were classically identified within anaerobic treatment systems and expected with known functions, a substantial fraction of the clone sequences showed relatively low levels of similarity with any other reported rDNA sequences and thus were derived from unknown taxa. These results suggest that bacterial communities in landfill environment are far more complex than previously expected and remain largely unexplored.

Bacteria↗
PubMed2004

Quantifying intrachromosomal GC heterogeneity in prokaryotic genomes.

The sequencing of prokaryotic genomes covering a wide taxonomic range has sparked renewed interest in intrachromosomal compositional (GC) heterogeneity, largely in view of lateral transfers. We present here a brief overview of some methods for visualizing and quantifying GC variation in prokaryotes. We used these methods to examine heterogeneity levels in sequenced prokaryotes, for a range of scales or stringencies. Some species are consistently homogeneous, whereas others are markedly heterogeneous in comparison, in particular Aeropyrum pernix, Xylella fastidiosa, Mycoplasma genitalium, Enterococcus faecalis, Bacillus subtilis, Pyrobaculum aerophilum, Vibrio vulnificus chromosome I, Deinococcus radiodurans chromosome II and Halobacterium. As we discuss here, the wide range of heterogeneities calls for reexamination of an accepted belief, namely that the endogenous DNA of bacteria and archaea should typically exhibit low intrachromosomal GC contrasts. Supplementary results for all species analyzed are available at our website: http://bioinfo2.ugr.es/prok.

Algorithms↗
PubMed2005

Characterization of heterologous hemoglobin and flavohemoglobin promoter regulation in Escherichia coli.

Bacterial hemoglobins and flavohemoglobins have been used to improve cell growth and productivity in biotechnological applications. The expression of globin genes can be induced by reducing the oxygen supply or applying external stressors, which provide a simple and inexpensive mechanism for induction of heterologous protein production. It is in the interest of the biotechnological industry to seek new promoters, which are non-patented, cheap and simple to induce. Therefore, new globin gene promoters have been isolated from Campylobacter jejuni, Bacillus subtilis, Deinococcus radiodurans, Streptomyces coelicolor, and Salmonella typhi. The goal was to obtain insights about the regulation mechanisms of these promoters in Escherichia coli using in silico and experimental methods. The recognition of these promoters by the E. coli transcriptional machinery was first analyzed by computational methods. Computer analysis revealed that all the promoters, except the promoter of S. coelicolor, should be functional in E. coli and most of them also contain putative binding sites for ArcA, CRP, and FNR global regulators. Furthermore, the expression profiles of the promoters fused to the chloramphenicol acetyl transferase gene were analyzed under various conditions using E. coli mutants devoid of regulatory molecules. In vivo regulation studies of globin promoters mainly verified the in silico predictions.

Bacteria↗
PubMed2006

Structural insights into enzyme-substrate interaction and characterization of enzymatic intermediates of organic hydroperoxide resistance protein from Xylella fastidiosa.

Organic hydroperoxide resistance proteins (Ohr) belong to a family of proteins that possess thiol-dependent peroxidase activity endowed by reactive cysteine residues able to reduce peroxides. The crystal structure of Ohr from Xylella fastidiosa in complex with polyethylene glycol, providing insights into enzyme-substrate interactions is described herein. In addition, crystallographic studies, molecular modeling and biochemical assays also indicated that peroxides derived from long chain fatty acids could be the biological substrates of Ohr. Because different oxidation states of the reactive cysteine were present in the Ohr structures from X. fastidiosa, Pseudomonas aeruginosa and Deinococcus radiodurans it was possible to envisage a set of snapshots along the coordinate of the enzyme-catalyzed reaction. The redox intermediates of X. fastidiosa Ohr observed in the crystals were further characterized in solution by electrospray ionization mass spectrometry and by biochemical approaches. In this study, the formation of an intramolecular disulfide bond and oxidative inactivation through the formation of a sulfonic acid derivative was unequivocally demonstrated for the first time. Because Ohr proteins are exclusively present in bacteria, they may represent promising targets for therapeutical drugs. In this regard, the structural and functional analyses of Ohr presented here might be very useful.

Animals↗