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Meiothermus taiwanensis sp. nov., a novel filamentous, thermophilic species isolated in Taiwan.

Two novel filamentous bacterial isolates, strains WR-30T and WR-220, with an optimum growth temperature of approximately 55-60 degrees C were isolated from Wu-rai hot springs in the northern part of Taiwan. These isolates were aerobic, thermophilic, non-sporulating, red-pigmented and heterotrophic and formed extremely long, filamentous trichomes from cells of different lengths. Phylogenetic analysis of 16S rDNA, DNA-DNA hybridization, morphological and biochemical features and fatty acid composition revealed that the isolates represent a novel species of the genus Meiothermus. The name Meiothermus taiwanensis sp. nov. is proposed for this novel species. The type strain of M. taiwanensis is strain WR-30T (= ATCC BAA-399T = CCRC 17170T = DSM 14542T); strain WR-220 (= ATCC BAA400 = CCRC 17171 = DSM 14543) is a reference strain.

Base Composition↗

Reproducible growth of well diffracting ribosomal crystals.

The crystallization of ribosomal particles is associated with extraordinary challenging demands. This originates mainly from the ribosome's natural tendency to deteriorate and from its multi-conformational heterogeneity, both of which stem from its functional flexibility. To increase the level of homogeneity of ribosomal preparations, systematic searches for conditions yielding populations of fully defined chemical compositions were employed and the variables essential for high functional activity were analyzed and optimized. These include temperature, cell-growth duration and media, the cell-harvesting stage, ribosomal purification and storage. The functional state that is most suitable to yield quality crystals was identified as that of the polysome and it was found that this fraction reproducibly yielded crystals of superior properties.

Crystallography, X-Ray↗

Is radiation damage dependent on the dose rate used during macromolecular crystallography data collection?

This paper focuses on the radiation-damage effects when applying the same total X-ray dose to protein crystals at different dose rates. These experiments have been performed on both a selenomethionated protein and on bovine trypsin using dose rates that span nearly two orders of magnitude. The results show no clear dose-rate effect on the global indicators of radiation damage, but a small measurable dose-rate effect could be found when studying specific radiation damage. It is hypothesized that this observed dose-rate effect relates to differences in the steady-state free-radical concentration.

Animals↗

Structural determination of the polar glycoglycerolipids from thermophilic bacteria Meiothermus taiwanensis.

The polar glycolipids were isolated from the thermophilic bacteria Meiothermus taiwanensis ATCC BAA-400 by ethanol extraction and purified by Sephadex LH-20 and silica gel column chromatography. The fatty acid composition of O-acyl groups in the glycolipids was obtained by gas chromatography mass spectroscopy analysis on their methyl esters derived from methanolysis and was made mainly of C(15:0) (34.0%) and C(17:0) (42.3%) fatty acids, with the majority as branched fatty acids (over 80%). Removal of O-acyl groups under mild basic conditions provided two glycolipids, which differ only in N-acyl substitution on a hexosamine. Electrospray mass spectroscopy analysis revealed that one has a C(17:0) N-acyl group and the other hydroxy C(17:0) in a ratio of about 1 : 3.5. Furthermore, complete de-lipidation with strong base followed by selective N-acetylation resulted in a homogeneous tetraglycosyl glycerol. The linkages and configurations of the carbohydrate moiety were then elucidated by MS and various NMR analyses. Thus, the major glycolipid from M. taiwanensis ATCC BAA-400 was determined with the following structure: alpha-Galp(1-6)-beta-Galp(1-6)-beta-GalNAcyl(1,2)-alpha-Glc(1,1)-Gro diester, where N-acyl is C(17:0) or hydroxy C(17:0) fatty acid and the glycerol esters were mainly iso- and anteisobranched C(15:0) and C(17:0).

Acylation↗

INACTIVATION OF THE RADIATION-RESISTANT SPOILAGE BACTERIUM MICROCOCCUS RADIODURANS. I. RADIATION INACTIVATION RATES IN THREE MEAT SUBSTRATES AND IN BUFFER.

A simplified technique permitting the pipetting of raw puréed meats for quantitative bacteriological study is described for use in determining survival of these non-sporing bacteria, which are exceptionally resistant to radiation. Survival curves, using gamma radiation as the sterilizing agent, were determined in raw beef with four strains of Micrococcus radiodurans. Survival curves of the R(1) strain in other meat substrates showed that survival was significantly greater in raw beef and raw chicken than in raw fish or in cooked beef. Resistance was lowest in the buffer. Cells grown in broth (an artificial growth medium) and resuspended in beef did not differ in resistance from cells that had been grown and irradiated in beef. Survival rate was statistically independent of the initial cell concentration, even though there appeared to be a correlation between lower death rate and lower initial cell concentrations. The initial viable count of this culture of the domesticated R(1) strain in beef was reduced by a factor of about 10(-5) by 3.0 megarad, and 4.0 megarad reduced the initial count by a factor of more than 10(-9). Data suggest that M. radiodurans R(1) is more resistant to radiation than spore-forming spoilage bacteria for which inactivation rates have been published.

Animals↗

INACTIVATION OF THE RADIATION-RESISTANT SPOILAGE BACTERIUM MICROCOCCUS RADIODURANS. II. RADIATION INACTIVATION RATES AS INFLUENCED BY MENSTRUUM TEMPERATURE, PREIRRADIATION HEAT TREATMENT, AND CERTAIN REDUCING AGENTS.

The R(1) strain of Micrococcus radiodurans, previously determined to be more resistant than three other strains exposed to gamma radiation, was studied further to determine the influence of certain environmental factors on resistance to radiation inactivation. The frozen state offered insignificant protection to the organisms irradiated in raw puréed beef. Resistance was reduced by higher menstruum temperatures (40 and 50 C) during irradiation. Preirradiation heat treatment was found to lower resistance to subsequent irradiation. When the cells were irradiated in buffer at pH 5, 7, or 9, no differences in resistance were noted. Cell suspensions in buffer were protected to some extent by cysteine but not by thioglycolate. Ascorbate enhanced radiation inactivation.

Animals↗

Antibiotics targeting ribosomes: resistance, selectivity, synergism and cellular regulation.

Antibiotics target ribosomes at distinct locations within functionally relevant sites. They exert their inhibitory action by diverse modes, including competing with substrate binding, interfering with ribosomal dynamics, minimizing ribosomal mobility, facilitating miscoding, hampering the progression of the mRNA chain, and blocking the nascent protein exit tunnel. Although the ribosomes are highly conserved organelles, they possess subtle sequence and/or conformational variations. These enable drug selectivity, thus facilitating clinical usage. The structural implications of these differences were deciphered by comparisons of high-resolution structures of complexes of antibiotics with ribosomal particles from eubacteria resembling pathogens and from an archaeon that shares properties with eukaryotes. The various antibiotic-binding modes detected in these structures demonstrate that members of antibiotic families possessing common chemical elements with minute differences might bind to ribosomal pockets in significantly different modes, governed by their chemical properties. Similarly, the nature of seemingly identical mechanisms of drug resistance is dominated, directly or via cellular effects, by the antibiotics' chemical properties. The observed variability in antibiotic binding and inhibitory modes justifies expectations for structurally based improved properties of existing compounds as well as for the discovery of novel drug classes.

Anti-Bacterial Agents↗

A rational approach for discovering and validating cancer markers in very small samples using mass spectrometry and ELISA microarrays.

Identifying useful markers of cancer can be problematic due to limited amounts of sample. Some samples such as nipple aspirate fluid (NAF) or early-stage tumors are inherently small. Other samples such as serum are collected in larger volumes but archives of these samples are very valuable and only small amounts of each sample may be available for a single study. Also, given the diverse nature of cancer and the inherent variability in individual protein levels, it seems likely that the best approach to screen for cancer will be to determine the profile of a battery of proteins. As a result, a major challenge in identifying protein markers of disease is the ability to screen many proteins using very small amounts of sample. In this review, we outline some technological advances in proteomics that greatly advance this capability. Specifically, we propose a strategy for identifying markers of breast cancer in NAF that utilizes mass spectrometry (MS) to simultaneously screen hundreds or thousands of proteins in each sample. The best potential markers identified by the MS analysis can then be extensively characterized using an ELISA microarray assay. Because the microarray analysis is quantitative and large numbers of samples can be efficiently analyzed, this approach offers the ability to rapidly assess a battery of selected proteins in a manner that is directly relevant to traditional clinical assays.

Bacterial Proteins↗

The TROVE module: a common element in Telomerase, Ro and Vault ribonucleoproteins.

BACKGROUND: Ribonucleoproteins carry out a variety of important tasks in the cell. In this study we show that a number of these contain a novel module, that we speculate mediates RNA-binding. RESULTS: The TROVE module--Telomerase, Ro and Vault module--is found in TEP1 and Ro60 the protein components of three ribonucleoprotein particles. This novel module, consisting of one or more domains, may be involved in binding the RNA components of the three RNPs, which are telomerase RNA, Y RNA and vault RNA. A second conserved region in these proteins is shown to be a member of the vWA domain family. The vWA domain in TEP1 is closely related to the previously recognised vWA domain in VPARP a second component of the vault particle. This vWA domain may mediate interactions between these vault components or bind as yet unidentified components of the RNPs. CONCLUSIONS: This work suggests that a number of ribonucleoprotein components use a common RNA-binding module. The TROVE module is also found in bacterial ribonucleoproteins suggesting an ancient origin for these ribonucleoproteins.

Amino Acid Motifs↗

Gene prediction using the Self-Organizing Map: automatic generation of multiple gene models.

BACKGROUND: Many current gene prediction methods use only one model to represent protein-coding regions in a genome, and so are less likely to predict the location of genes that have an atypical sequence composition. It is likely that future improvements in gene finding will involve the development of methods that can adequately deal with intra-genomic compositional variation. RESULTS: This work explores a new approach to gene-prediction, based on the Self-Organizing Map, which has the ability to automatically identify multiple gene models within a genome. The current implementation, named RescueNet, uses relative synonymous codon usage as the indicator of protein-coding potential. CONCLUSIONS: While its raw accuracy rate can be less than other methods, RescueNet consistently identifies some genes that other methods do not, and should therefore be of interest to gene-prediction software developers and genome annotation teams alike. RescueNet is recommended for use in conjunction with, or as a complement to, other gene prediction methods.

Chromosome Mapping↗

Asymmetric directional mutation pressures in bacteria.

BACKGROUND: When there are no strand-specific biases in mutation and selection rates (that is, in the substitution rates) between the two strands of DNA, the average nucleotide composition is theoretically expected to be A = T and G = C within each strand. Deviations from these equalities are therefore evidence for an asymmetry in selection and/or mutation between the two strands. By focusing on weakly selected regions that could be oriented with respect to replication in 43 out of 51 completely sequenced bacterial chromosomes, we have been able to detect asymmetric directional mutation pressures. RESULTS: Most of the 43 chromosomes were found to be relatively enriched in G over C and T over A, and slightly depleted in G+C, in their weakly selected positions (intergenic regions and third codon positions) in the leading strand compared with the lagging strand. Deviations from A = T and G = C were highly correlated between third codon positions and intergenic regions, with a lower degree of deviation in intergenic regions, and were not correlated with overall genomic G+C content. CONCLUSIONS: During the course of bacterial chromosome evolution, the effects of asymmetric directional mutation pressures are commonly observed in weakly selected positions. The degree of deviation from equality is highly variable among species, and within species is higher in third codon positions than in intergenic regions. The orientation of these effects is almost universal and is compatible in most cases with the hypothesis of an excess of cytosine deamination in the single-stranded state during DNA replication. However, the variation in G+C content between species is influenced by factors other than asymmetric mutation pressure.

AT Rich Sequence↗

Evolution of mosaic operons by horizontal gene transfer and gene displacement in situ.

BACKGROUND: Shuffling and disruption of operons and horizontal gene transfer are major contributions to the new, dynamic view of prokaryotic evolution. Under the 'selfish operon' hypothesis, operons are viewed as mobile genetic entities that are constantly disseminated via horizontal gene transfer, although their retention could be favored by the advantage of coregulation of functionally linked genes. Here we apply comparative genomics and phylogenetic analysis to examine horizontal transfer of entire operons versus displacement of individual genes within operons by horizontally acquired orthologs and independent assembly of the same or similar operons from genes with different phylogenetic affinities. RESULTS: Since a substantial number of operons have been identified experimentally in only a few model bacteria, evolutionarily conserved gene strings were analyzed as surrogates of operons. The phylogenetic affinities within these predicted operons were assessed first by sequence similarity analysis and then by phylogenetic analysis, including statistical tests of tree topology. Numerous cases of apparent horizontal transfer of entire operons were detected. However, it was shown that apparent horizontal transfer of individual genes or arrays of genes within operons is not uncommon either and results in xenologous gene displacement in situ, that is, displacement of an ancestral gene by a horizontally transferred ortholog from a taxonomically distant organism without change of the local gene organization. On rarer occasions, operons might have evolved via independent assembly, in part from horizontally acquired genes. CONCLUSIONS: The discovery of in situ gene displacement shows that combination of rampant horizontal gene transfer with selection for preservation of operon structure provides for events in prokaryotic evolution that, a priori, seem improbable. These findings also emphasize that not all aspects of operon evolution are selfish, with operon integrity maintained by purifying selection at the organism level.

Alkyl and Aryl Transferases↗