Kineococcus Radiotolerans Research Articles

Isolation and Molecular Diagnosis of Enterobacter cloacae and Kineococcus radiotolerans from Red Clover nodules and evaluation of the prepared inoculum from them as a stimulator for plant growth

To obtain biological inoculum that promotes plant growth and contributes to a clean environment and sustainable agriculture, twentyone samples were collected from the root nodes of the red clover plant (Trifolium paratens) grown in gypsiferous soils at the Research Station of the Department of Soil Sciences and Water Resources - Faculty of Agriculture - University of Tikrit, samples were cultured in the medium of YEMA. Two isolates belonging to Enterobacter cloacae and Kineococcus radiotolerans were diagnosed phenotypically and molecularly. The diagnosis was confirmed by analyzing the nitrogen bases sequence of the DNA of the 16S rRNA gene. It was recorded in the National Center Biotechnology Information(NCBI) with the numbers MN310027.1 and KT216573.1 respectively, these isolates were activated and tested their efficiency as a bio-stimulant by testing their ability to dissolve insoluble phosphate compounds and produce indole acetic acid and chelating compounds, the results showed the ability of all isolates to produce indole acetic acid (IAA), chelating compounds, and solubility of phosphates, E. cloacae isolate outperformed K. radiotolerans isolates in these parameters, as it gave E. coloaca solubilization of phosphate and IAA production of 40.3 mg p L-1 and 11.2 µg ml-1 respectively, and it showed a high output of the chelating compounds compared to the treatment inoculated with K. radiotolerans, which gave 27.20 mg p L-1 and 7.21 µg ml-1, and medium production of iron chelating compounds, the results also showed the superiority of the inoculated treatments over the uninoculated treatments in the percentage of germination, the speed of germination, the length of tomato seedlings, the dry weight of the Shoot and root parts and the number of leaves, and the results showed the superiority of the inoculated treatment with E. cloacae significantly on K. radiotolerans isolate. Keywords: Enterobacter cloacae; Kineococcus radiotolerans; Molecular Diagnosis; Bacterial inoculation; Eggplant seedlings.

TOMATO SEEDLING PRODUCTION USING AN INOCULUM PREPARED WITH PLANT GROWTH-PROMOTING RHIZOBACTERIA (PGPR) ISOLATES

A study to attain healthy tomato (Lycopersicon esculentum Mill) seedlings through environment-friendly natural biological products took place from January to March 2022. Determining the effects of the inoculum prepared from five plant growth-promoting rhizobacteria (PGPR) isolates, i.e., Pseudomonas fluorescens, Pseudomonas putida, Lysinibacillus fusiformis, Enterobacter cloacae, and Kineococcus radiotolerans on the production of tomato seedlings, experiments proceeded at the Soil Microbiology Laboratory, Department of Soil Science and Water Resources, College of Agriculture, Tikrit University, Iraq. The isolates underwent screening for their efficiency as a biostimulant to dissolve insoluble phosphate compounds and produce indole acetic acid (IAA) and chelating compounds. The results showed the ability of all the isolates to produce IAA, chelating compounds, and solubility of phosphates. The P. fluorescens isolate showed superior in its phosphate solubilization and IAA production (41.30 mg p-1, 13.00 mg ml-1), followed by P. putida, E. cloacae, L. fusiformis, and K. radiotolerans, respectively, with the production of medium chelating compounds. The results also showed the superiority of the inoculated treatments over the non-inoculated treatments in the percentage and speed of germination, the length of tomato seedlings, the shoot dry weight, the number of leaves per plant, and root weight parts. The treatments with P. fluorescens displayed significant superiority in all studied traits, followed by P. putida, E. cloacae, L. fusiformis, and K. radiotolerans.

Kineococcus vitellinus sp. nov., Kineococcus indalonis sp. nov. and Kineococcus siccus sp. nov., Isolated Nearby the Tabernas Desert (Almería, Spain)

Three novel Gram-positive, aerobic, chemoheterotrophic, motile, non-endospore-forming, orange-pigmented bacteria designated strains T13T, T90T and R8T were isolated from the Tabernas Desert biocrust (Almería, Spain). Cells of the three strains were coccus-shaped and occurred singly, in pairs or clusters. The three strains were oxidase-negative and catalase-positive, and showed a mesophilic, neutrophilic and non-halophilic metabolism. Based on the 16S rRNA gene sequences, the closest neighbours of strains T13T, T90T and R8T were Kineococcus aurantiacus IFO 15268T, Kineococcus gypseus YIM 121300T and Kineococcus radiotolerans SRS 30216T (98.5%, 97.1% and 97.9% gene sequence similarity, respectively). The genomes were sequenced, and have been deposited in the GenBank/EMBL/DDBJ databases under the accession numbers JAAALL000000000, JAAALM000000000 and JAAALN000000000, respectively, for strains T13T, T90T and R8T. The average nucleotide identity (ANIb) and digital DNA-DNA hybridization (dDDH) values confirmed their adscription to three new species within the genus Kineococcus. The genomic G + C content of strains T13T, T90T and R8T ranged from 75.1% to 76.3%. The predominant fatty acid of all three strains was anteiso-C15:0. According to a polyphasic study, strains T13T, T90T and R8T are representatives of three new species in the genus Kineococcus, for which names Kineococcus vitellinus sp. nov. (type strain T13T = CECT 9936T = DSM 110024T), Kineococcus indalonis sp. nov. (type strain T90T = CECT 9938T = DSM 110026T) and Kineococcus siccus sp. nov. (type strain R8T = CECT 9937T = DSM 110025T) are proposed.

Description of Chloramphenicol Resistant Kineococcus rubinsiae sp. nov. Isolated From a Spacecraft Assembly Facility.

A Gram-positive, coccoid, motile, aerobic bacterium, designated strain B12T was isolated from a Jet Propulsion Laboratory spacecraft assembly cleanroom, Pasadena, CA, United States. Strain B12T was resistant to chloramphenicol (100 μg/mL), and is a relatively slow grower (3–5 days optimal). Strain B12T was found to grow optimally at 28 to 32°C, pH 7 to 8, and 0.5% NaCl. Fatty acid methyl ester analysis showed that the major fatty acid of the strain B12T was anteiso C15:0 (66.3%), which is also produced by other Kineococcus species. However, arachidonic acid (C20:4 ω6,9,12,16c) was present in strain B12T and Kineococcus glutinatus YIM 75677T but absent in all other Kineococcus species. 16S rRNA analysis revealed that strain B12T was 97.9% similar to Kineococcus radiotolerans and falls within the Kineococcus clade. Low 16S rRNA gene sequence similarities (<94%) with other genera in the family Kineosporiaceae, including Angustibacter (93%), Kineosporia (94% to 95%), Pseudokineococcus (93%), Quadrisphaera (93%), and Thalassiella (94%) demonstrated that the strain B12T does not belong to these genera. Phylogenetic analysis of the gyrB gene show that all known Kineococcus species exhibited <86% sequence similarity with B12T. Multi-locus sequence and whole genome sequence analyses confirmed that B12T clades with other Kineococcus species. Average nucleotide identity of strain B12T were 75–78% with other Kineococcus species, while values ranged from 72–75% with species from other genera within family Kineosporiaceae. Average amino-acid identities were 66–72% with other Kineococcus species, while they ranged from 50–58% with species from other genera. The dDDH comparison of strain B12T genome with members of genera Kineococcus showed 20–22% similarity, again demonstrating that B12T is distantly related to other members of the genus. Furthermore, analysis of whole proteome deduced from WGS places strain B12T in order Kineosporiales, confirming that strain B12T is a novel member of family Kineosporiaceae. Based on these analyses and other genome characteristics, strain B12T is assigned to a novel species within the genus Kineococcus, and the name Kineococcus rubinsiae sp. nov., is proposed. The type strain is B12T (=FJII-L1-CM-PAB2T; NRRL B-65556T, DSM 110506T).

Metal-citrate complex transport in Kineococcus radiotolerans.

The growth of an organism is highly dependent on the acquisition of carbon and metals, and availability of these nutrients in the environment affects its survival. Organisms can obtain both nutrients simultaneously through proteins of the CitMHS superfamily. Bioinformatic studies suggested a CitMHS gene (Accession number ABS03965.1) in Kineococcus radiotolerans. Radio flux assays following 14-C radiolabelled citrate, either free or complexed to a variety of metal ions, in K. radiotolerans demonstrated internalization of the citrate when bound to select metal ions only, primarily in the form of calcium-citrate. A pH response was also observed, consistent with a permease (ATP independent) mechanism as noted for other CitMHS family members, with greater uptake at pH 7 compared to pH 10. These results confirm the ability of K. radiotolerans to transport complexed citrate.

Kineococcus terrestris sp. nov. and Kineococcus aureolus sp. nov., isolated from saline sediment.

Two novel actinobacteria, designated YIM 121936T and YIM 121940T, were isolated from alkaline sediment in Yuanjiang, China. The cells of the novel strains were Gram-stain-positive, aerobic, motile, non-spore-forming and coccus-shaped. The two strains both contained meso-diaminopimelic acid as the diagnostic diamino acid. The whole-cell sugars were arabinose, galactose, glucose, mannose and ribose. The predominant menaquinone was MK-9(H2). The polar lipid profile of both strains comprised diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannoside, one unknown phosphoglycolipid and five unknown phospholipids. The predominant fatty acids of both strains were anteiso-C15 : 0. The genomic DNA G+C contents of strains YIM 121936T and YIM 121940T were 74.7 and 75.2 %, respectively. Strain YIM 121936T was closely related to Kineococcus aurantiacus IFO 15268T (97.19 %), Kineococcus gypseus YIM 121300T (97.00 %) and Kineococcus mangrovi NBRC 110933T (97.00 %). Strain YIM 121940T was closely related to K. aurantiacus IFO 15268T (97.41 %), Kineococcus endophytica KLBMP 1274T (97.18 %), Kineococcus rhizosphaerae RP-B16T (97.09 %), Kineococcus radiotolerans SRS 30216T (97.09 %), K. gypseus YIM 121300T (97.00 %) and K. mangrovi NBRC 110933T (97.00 %). Strain YIM 121936T shared high 16S rRNA gene sequence similarity (99 %) with YIM 121940T. Similarities of two strains with other species of the genus Kineococcus were <97 %. The DNA-DNA hybridization values were below 70 % among all the strains studied. YIM 121936T and YIM 121940T are representatives of two new species in the genus Kineococcus, for which names Kineococcus terreus sp. nov. (type strain YIM 121936T=KCTC 39738T=DSM 102155T) and Kineococcus aureolus sp. nov. (type strain YIM 121940T=KCTC 39739T=DSM 102158T) are proposed, respectively.

Transcriptome sequencing analysis of novel sRNAs of Kineococcus radiotolerans in response to ionizing radiation

Kineococcus radiotolerans is a Gram-positive, radio-resistant bacterium isolated from a radioactive environment. The small noncoding RNAs (sRNAs) in bacteria are reported to play roles in the immediate response to stress and/or the recovery from stress. The analysis of K. radiotolerans transcriptome sequencing results can identify these sRNAs in a genome-wide detection, using RNA sequencing (RNA-seq) by the deep sequencing technique. In this study, the raw data of radiation-exposed samples (RS) and control samples (CS) were acquired separately from the sequencing platform. There were 217 common sRNA candidates in the two samples screened in the genome-wide scale by bioinformatics analysis. There were 43 differentially expressed sRNA candidates, including 28 up-regulated and 15 down-regulated ones. The down-regulated sRNAs were selected for the sRNA target prediction, of which 12 sRNAs that may modulate the genes related to the transcription regulation and DNA repair were considered as the candidates involved in the radio-resistance regulation system.

Characterisation of caecum and crop microbiota of Indian indigenous chicken targeting multiple hypervariable regions within 16S rRNA gene

A comparative analysis of caecum and crop microbiota of chick, grower and adult stages of Indian indigenous chickens was conducted to investigate the role of the microbiota of the gastrointestinal tract, which play an important role in host performance, health and immunity.High-throughput Illumina sequencing was performed for V3, V4 and V4-V6 hypervariable regions of the 16S rRNA gene. M5RNA and M5NR databases under MG-RAST were used for metagenomic datasets annotation.In the crop, Firmicutes (~78%) and Proteobacteria (~16%) were the predominant phyla whereas in the caecum, Firmicutes (~50%), Bacteroidetes (~29%) and Actinobacteria (~10%) were predominant. The Shannon-Wiener diversity index suggested that sample richness and diversity increased as the chicken aged.For the first time, the presence of Lactobacillus species such as L. frumenti, L. antri, L. mucosae in the chicken crop along with Kineococcus radiotolerans, Desulfohalobium retbaense and L. jensenii in the caecum are reported. Many of these bacterial species have been found to be involved in immune response modulation and disease prevention in pigs and humans. The gut microbiome of the indigenous chicken was enriched with microbes having probiotic potential which might be essential for their adaptability.

Novel genomic island modifies DNA with 7-deazaguanine derivatives

The discovery of ∼20-kb gene clusters containing a family of paralogs of tRNA guanosine transglycosylase genes, called tgtA5, alongside 7-cyano-7-deazaguanine (preQ0) synthesis and DNA metabolism genes, led to the hypothesis that 7-deazaguanine derivatives are inserted in DNA. This was established by detecting 2'-deoxy-preQ0 and 2'-deoxy-7-amido-7-deazaguanosine in enzymatic hydrolysates of DNA extracted from the pathogenic, Gram-negative bacteria Salmonella enterica serovar Montevideo. These modifications were absent in the closely related S. enterica serovar Typhimurium LT2 and from a mutant of S Montevideo, each lacking the gene cluster. This led us to rename the genes of the S. Montevideo cluster as dpdA-K for 7-deazapurine in DNA. Similar gene clusters were analyzed in ∼150 phylogenetically diverse bacteria, and the modifications were detected in DNA from other organisms containing these clusters, including Kineococcus radiotolerans, Comamonas testosteroni, and Sphingopyxis alaskensis Comparative genomic analysis shows that, in Enterobacteriaceae, the cluster is a genomic island integrated at the leuX locus, and the phylogenetic analysis of the TgtA5 family is consistent with widespread horizontal gene transfer. Comparison of transformation efficiencies of modified or unmodified plasmids into isogenic S. Montevideo strains containing or lacking the cluster strongly suggests a restriction-modification role for the cluster in Enterobacteriaceae. Another preQ0 derivative, 2'-deoxy-7-formamidino-7-deazaguanosine, was found in the Escherichia coli bacteriophage 9 g, as predicted from the presence of homologs of genes involved in the synthesis of the archaeosine tRNA modification. These results illustrate a deep and unexpected evolutionary connection between DNA and tRNA metabolism.

Deep sequencing analysis of the Kineococcus radiotolerans transcriptome in response to ionizing radiation

Kineococcus radiotolerans is a gram-positive, radiation-resistant bacterium that was isolated from a radioactive environment. The synergy of several groups of genes is thought to contribute to the radio-resistance of this species of bacteria. Sequencing of the transcriptome, RNA sequencing (RNA-seq), using deep sequencing technology can reveal the genes that are differentially expressed in response to radiation in this bacterial strain. In this study, the transcriptomes of two samples (with and without irradiation treatment) were sequencing by deep sequencing technology. After the bioinformatics process, 143 genes were screened out by the differential expression (DE) analysis. In all 143 differentially expressed genes, 20 genes were annotated to be related to the radio-resistance based on the cluster analysis by the cluster of orthologous groups of proteins (COG) annotation which were validated by the quantitative RT-PCR. The pathway analysis revealed that these 20 validated genes were related to DNA damage repair, including recA, ruvA and ruvB, which were considered to be the key genes in DNA damage repair. This study provides the foundation to investigate the regulatory mechanism of these genes.

Kineococcus radiotolerans Dps forms a heteronuclear Mn–Fe ferroxidase center that may explain the Mn-dependent protection against oxidative stress

BackgroundThe ferroxidase center of DNA-binding protein from starved cells (Dps) is a major player in the iron oxidation/detoxification process that leads to a decreased reactive oxygen species production. The possible Mn(II) participation in this process has been studied in Dps from Kineococcus radiotolerans, a radiation-resistant bacterium with a high cytosolic Mn/Fe ratio and a high capacity to survive ionizing and stress conditions. MethodsThe X-ray structure of recombinant K. radiotolerans Dps loaded with Mn(II) has been solved at 2.0Å resolution. Mn(II) binding to K. radiotolerans Dps and its effect on Fe(II) oxidation have been characterized in spectroscopic measurements. ResultsIn K. radiotolerans Dps, the Fe–Fe ferroxidase center can have a Mn–Fe composition. Mn(II) binds only at the high affinity, so-called A site, whereas Fe(II) binds also at the low affinity, so-called B site. The Mn–Fe and Fe–Fe centers behave distinctly upon iron oxidation by O2. A site-bound Mn(II) or Fe(II) plays a catalytic role, while B site-bound Fe(II) behaves like a substrate and can be replaced by another Fe(II) after oxidation. When H2O2 is the Fe(II) oxidant, single electrons are transferred to aromatic residues near the ferroxidase center and give rise to intra-protein radicals thereby limiting OH release in solution. The presence of the Mn–Fe center results in significant differences in the development of such intra-protein radicals. ConclusionsMn(II) bound at the Dps ferroxidase center A site undergoes redox cycling provided the B site contains Fe. General significanceThe results provide a likely molecular mechanism for the protective role of Mn(II) under oxidative stress conditions as it participates in redox cycling in the hetero-binuclear ferroxidase center.

Kineococcus endophytica sp. nov., a novel endophytic actinomycete isolated from a coastal halophyte in Jiangsu, China

A novel Gram-positive, motile, non-spore-forming coccus-shaped bacterial strain, designated KLBMP 1274(T), was isolated from a halophytic plant (Limonium sinense) collected from the coastal region of Nantong, Jiangsu Province, in east China. Phylogenetic analyses based on the 16S rRNA gene sequence showed that strain KLBMP 1274(T) belongs to the genus Kineococcus and is closely related to Kineococcus rhizosphaerae RP-B16(T) (98.72%), Kineococcus aurantiacus IFO 15268(T) (98.71%), Kineococcus radiotolerans SRS30216(T) (98.69%) and Kineococcus gynurae KKD096(T) (97.33%). The 16S rRNA gene sequence similarity to other species of the genus Kineococcus was <97%. The cell wall contained meso-diaminopimelic acid as the diagnostic diamino acid, with arabinose and galactose as the characteristic sugars. The predominant menaquinone was MK-9(H(2)). The polar lipids were found to be diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannosides, an unknown phospholipid, an unknown glycolipid, and three unknown lipids. Major cellular fatty acids were found to be anteiso-C(15: 0) and iso-C(14: 0). The chemotaxonomic data for strain KLBMP 1274(T) were typical of the genus Kineococcus. The total DNA G+C content was 73.4mol%. DNA-DNA relatedness and differential phenotypic data demonstrated that strain KLBMP 1274(T) was clearly distinguished from all closely related species of the genus Kineococcus. Thus, strain KLBMP 1274(T) represents a novel species of the genus Kineococcus, for which the name Kineococcus endophytica sp. nov. is proposed. The type strain is KLBMP 1274(T) (=KCTC 19886 (T)=NBRC 108674(T)).

Proteomic and Physiological Responses of Kineococcus radiotolerans to Copper

Copper is a highly reactive, toxic metal; consequently, transport of this metal within the cell is tightly regulated. Intriguingly, the actinobacterium Kineococcus radiotolerans has been shown to not only accumulate soluble copper to high levels within the cytoplasm, but the phenotype also correlated with enhanced cell growth during chronic exposure to ionizing radiation. This study offers a first glimpse into the physiological and proteomic responses of K. radiotolerans to copper at increasing concentration and distinct growth phases. Aerobic growth rates and biomass yields were similar over a range of Cu(II) concentrations (0–1.5 mM) in complex medium. Copper uptake coincided with active cell growth and intracellular accumulation was positively correlated with Cu(II) concentration in the growth medium (R2 = 0.7). Approximately 40% of protein coding ORFs on the K. radiotolerans genome were differentially expressed in response to the copper treatments imposed. Copper accumulation coincided with increased abundance of proteins involved in oxidative stress and defense, DNA stabilization and repair, and protein turnover. Interestingly, the specific activity of superoxide dismutase was repressed by low to moderate concentrations of copper during exponential growth, and activity was unresponsive to perturbation with paraquot. The biochemical response pathways invoked by sub-lethal copper concentrations are exceptionally complex; though integral cellular functions are preserved, in part, through the coordination of defense enzymes, chaperones, antioxidants and protective osmolytes that likely help maintain cellular redox. This study extends our understanding of the ecology and physiology of this unique actinobacterium that could potentially inspire new biotechnologies in metal recovery and sequestration, and environmental restoration.

Novel blue light‐sensitive proteins from a metagenomic approach

A microarray-based approach was used to screen a soil metagenome for the presence of blue light (BL) photoreceptor-encoding genes. The microarray carried 149 different 54-mer oligonucleotides, derived from consensus sequences of light, oxygen and voltage (LOV) domain BL photoreceptor genes. Calibration of the microarrays allowed the detection of minimally 50 ng of genomic DNA against a background of 2-5 microg of genomic DNA. Identification of a positive cosmid clone was still possible for an amount of 0.25 ng against a background of 10 microg of labelled DNA clones. The array could readily identify targets carrying 4% sequence mismatch. Using the LOV microarray, up to 1200 library clones in concentrations of c. 20 ng each with a c. 40 kb insert size could be screened in a single batch. After calibration and reliability controls, the microarray was probed with cosmid-cloned DNA from the thermophilic fraction of a soil sample. From this approach, a novel gene was isolated that encodes a protein consisting of several Per-Arnt-Sim domains, a LOV domain associated to a histidine kinase and a response regulator domain. The novel gene showed highest similarity to a known sequence from Kineococcus radiotolerans SRS30216 (58% identity for the LOV domain only) and to a gene from Methylibium petroleiphilum PM1 (57% identity). The gene, designated as ht-met1 (Hamburg Thermophile Metagenome 1), was isolated and fully sequenced (3615 bp). ht-met1 is followed by a second open reading frame encoding a Fe-chelatase, an arrangement quite frequent for BL photoreceptors. The LOV domain region of ht-met1 was subcloned and expressed yielding a fully functional, flavin-containing LOV domain. Irradiation generated the typical LOV photochemistry, with the transient formation of a flavin-protein photoadduct. The dark recovery lifetime was found as tau(REC) = 120 s (20 degrees C) and is among the fastest ones determined so far for bacterial LOV domains.

Kineococcus rhizosphaerae sp. nov., isolated from rhizosphere soil

An orange-coloured actinomycete, designated strain RP-B16T, was isolated from a rhizosphere soil. Cells of strain RP-B16T were strictly aerobic, Gram-positive-staining, oxidase-negative, catalase-positive, motile cocci. A phylogenetic tree, based on 16S rRNA gene sequences, showed that members of the genus Kineococcus were divided into two independent clusters and that strain RP-B16T formed a coherent clade with the type strains of Kineococcus aurantiacus (99.5% sequence similarity) and Kineococcus radiotolerans (99.1%). Levels of 16S rRNA gene sequence similarity between strain RP-B16T and Kineococcus marinus KST3-3T and 'Kineococcus tegulae' T2A-S27 were lower than 97.0%. Chemotaxonomic characteristics were typical of the genus Kineococcus in having meso-diaminopimelic acid as the diagnostic diamino acid, MK-9(H2) as the predominant menaquinone, a polar lipid profile comprising diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and two unknown phospholipids, anteiso-C15:0 as the predominant fatty acid and a DNA G+C content of 73.8 mol%. Strain RP-B16T showed levels of DNA-DNA relatedness of 34.6-37.3% to Kineococcus aurantiacus DSM 7487T and 31.2-32.7% to Kineococcus radiotolerans DSM 14245T. On the basis of phenotypic features and DNA-DNA hybridization data, strain RP-B16T is considered to represent a novel species of the genus Kineococcus, for which the name Kineococcus rhizosphaerae sp. nov. is proposed. The type strain is RP-B16T (=KCTC 19366T=DSM 19711T).

DNA-Dependent ATPase Activity of Bacterial XPB Helicases

XPB, the largest subunit of the eukaryotic transcription factor TFIIH, is essential for both initiation of transcription by RNA polymerase II and nucleotide excision repair (NER). XPB belongs to the SF2 superfamily of monomeric helicases. XPB helicase is thought to have evolved in eukaryotes; however, a gene highly homologous to human XPB can be found in a number of bacteria. This report is the first biochemical characterization of XPB homologues from bacteria, specifically those from Mycobacterium tuberculosis and Kineococcus radiotolerans. Similarly to eukaryotic XPB, bacterial XPB are ATP-dependent 3' --> 5' DNA helicases. The ATPase activity of these XPB helicases is DNA-dependent, requiring a minimum of 4-nucleotide long single-stranded DNA (ssDNA). The maximum rates of ATP hydrolysis are about 10 and 50 molecules per minute by one XPB monomer on a 21-nucleotide ssDNA oligomer and on 5-kb long circular ssDNA, respectively. The ATP hydrolysis by the bacterial XPBs is coupled to their translocation along single-stranded DNA. The hydrolytic activity is strongly dependent on both the nature of a nucleotide triphosphate and that of a divalent metal. The inefficient ATP hydrolysis by bacterial XPB is consistent with nonprocessive functions of its eukaryotic homologue in locally remodeling DNA during transcription initiation and NER.

Gene Expression Analysis of Four Radiation-resistant Bacteria.

To investigate the general radiation-resistant mechanisms of bacteria, bioinformatic method was employed to predict highly expressed genes for four radiation-resistant bacteria, i.e. Deinococcus geothermalis (D. geo), Deinococcus radiodurans (D. rad), Kineococcus radiotolerans (K. rad) and Rubrobacter xylanophilus (R. xyl). It is revealed that most of the three reference gene sets, i.e. ribosomal proteins, transcription factors and major chaperones, are generally highly expressed in the four bacteria. Recombinase A (recA), a key enzyme in recombinational repair, is predicted to be highly or marginally highly expressed in the four bacteria. However, most proteins associated with other repair systems show low expression levels. Some genes participating in 'information storage and processing,' 'cellular processes and signaling' and 'metabolism' are among the top twenty predicted highly expressed (PHX) genes in the four genomes. Many antioxidant enzymes and proteases are commonly highly expressed in the four bacteria, indicating that these enzymes play important roles in resisting irradiation. Finally, a number of 'hypothetical genes' are among the top twenty PHX genes in each genome, some of them might contribute vitally to resist irradiation. Some of the prediction results are supported by experimental evidence. All the above information not only helps to understand the radiation-resistant mechanisms but also provides clues for identifying new radiation-resistant genes from these bacteria.

Survival in Nuclear Waste, Extreme Resistance, and Potential Applications Gleaned from the Genome Sequence of Kineococcus radiotolerans SRS30216

Kineococcus radiotolerans SRS30216 was isolated from a high-level radioactive environment at the Savannah River Site (SRS) and exhibits γ-radiation resistance approaching that of Deinococcus radiodurans. The genome was sequenced by the U.S. Department of Energy's Joint Genome Institute which suggested the existence of three replicons, a 4.76 Mb linear chromosome, a 0.18 Mb linear plasmid, and a 12.92 Kb circular plasmid. Southern hybridization confirmed that the chromosome is linear. The K. radiotolerans genome sequence was examined to learn about the physiology of the organism with regard to ionizing radiation resistance, the potential for bioremediation of nuclear waste, and the dimorphic life cycle. K. radiotolerans may have a unique genetic toolbox for radiation protection as it lacks many of the genes known to confer radiation resistance in D. radiodurans. Additionally, genes involved in the detoxification of reactive oxygen species and the excision repair pathway are overrepresented. K. radiotolerans appears to lack degradation pathways for pervasive soil and groundwater pollutants. However, it can respire on two organic acids found in SRS high-level nuclear waste, formate and oxalate, which promote the survival of cells during prolonged periods of starvation. The dimorphic life cycle involves the production of motile zoospores. The flagellar biosynthesis genes are located on a motility island, though its regulation could not be fully discerned. These results highlight the remarkable ability of K radiotolerans to withstand environmental extremes and suggest that in situ bioremediation of organic complexants from high level radioactive waste may be feasible.

Kineococcus gynurae sp. nov., isolated from a Thai medicinal plant

A novel, Gram-positive, motile, coccus-shaped, orange-pigmented organism, designated strain KKD096(T), was isolated from the roots of a Thai medicinal plant, Gynura pseudochina DC. var. hispida Thwaites. Growth of strain KKD096(T) occurred at temperatures of 14-34 degrees C, at pH 5.0-9.0 and at NaCl concentrations up to 7 % (w/v). Whole-cell hydrolysates contained arabinose and galactose as the characteristic sugars. The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid. The glycan moiety of the murein contained acetyl residues. The predominant menaquinone was MK-9(H2); mycolic acids were not detected. The genomic DNA G+C content was 73.3 mol%. The major cellular fatty acid was anteiso-C(15 : 0) (81.42 % of the total). Strain KKD096(T) was assigned to the genus Kineococcus on the basis of 16S rRNA gene sequence analysis; it was most closely related to Kineococcus radiotolerans DSM 14245(T) (97.1 % similarity). DNA-DNA hybridization revealed 39.4 % relatedness between these two taxa. On the basis of the genotypic and phenotypic data presented, strain KKD096(T) is considered to represent a novel species of the genus Kineococcus, for which the name Kineococcus gynurae sp. nov. is proposed. The type strain is KKD096(T) (=TISTR 1856(T)=NRRL B-24568(T)=BCC 26245(T)=NBRC 103943(T)).

Crystal structure of an alkaline serine protease from Nesterenkonia sp. defines a novel family of secreted bacterial proteases

Crystal structure of an alkaline serine protease from Nesterenkonia sp. defines a novel family of secreted bacterial proteases Na Yang, Jie Nan, Erik Brostromer, Rajni Hatti-Kaul, and Xiao-Dong Su* 1 Shenzhen Graduate School of Peking University, Shenzhen 518055, China 2 National Laboratory of Protein Engineering and Plant Genetic Engineering, Peking University, Beijing 100871, China 3 Department of Biotechnology, Center for Chemistry and Chemical Engineering, Lund University, S-221 00 Lund, Sweden