High Sequence Similarity (high + sequence_similarity)

Distribution by Scientific Domains

Selected Abstracts

Development of species-specific PCR primers on rDNA for the identification of European Armillaria species

G. Sicoli
Summary Attempts to design species-specific PCR primers from six European Armillaria species in the ribosomal RNA genes are reported. Primers were developed on the basis of the nucleotide sequence variability of the internal transcribed spacers (ITS) and the intergenic spacer (IGS1) of the ribosomal DNA. Four sets of primers gave specific PCR products for Armillaria tabescens, Armillaria mellea and Armillaria ostoyae. However, due to the high sequence similarities between Armillaria borealis and Armillaria ostoyae and between Armillaria cepistipes and Armillaria gallica no species specific amplification was obtained for these taxa. Résumé Des essais ont été réalisés pour obtenir des amorces PCR spécifiques de 6 espèces européennes d'Armillaria dans les gènes de l'ARNr. Les amorces ont été développées sur la base de la variabilité de séquence nucléotidique dans les ITS et IGS (IGS1) de l'ADN ribosomal. Quatre couples d'amorces ont permis d'obtenir des produits PCR spécifiques pour A. tabescens, A. mellea et A. ostoyae. Cependant, compte tenu des très fortes similarités de séquence entre A. borealis et A. ostoyae, et entre A. cepistipes et A. gallica, il n'a pas été obtenu d'amplification spécifique pour ces taxons. Zusammenfassung Es wird über Versuche berichtet, artspezifische Primer für sechs europäische Armillariaarten in der Region der ribosomalen RNA-Gene zu entwickeln. Als Grundlage dafür diente die Variabilität der Nukleotidsequenzen der ITS- und der IGS 1-Region der ribosomalen DNA. Vier Primerpaare ergaben spezifische PCR-Produkte für A. tabescens, A. mellea und A. ostoyae. Dagegen wurden aufgrund der grossen Ähnlichkeit der Sequenzen von A. borealis und A. ostoyae sowie von A. cepistipes und A. gallica für diese Taxa keine artspezifischen Amplifikationsprodukte erhalten. [source]

CoagMDB: a database analysis of missense mutations within four conserved domains in five vitamin K,dependent coagulation serine proteases using a text-mining tool,

HUMAN MUTATION, Issue 3 2008
Rebecca E. Saunders
Abstract Central repositories of mutations that combine structural, sequence, and phenotypic information in related proteins will facilitate the diagnosis and molecular understanding of diseases associated with them. Coagulation involves the sequential activation of serine proteases and regulators in order to yield stable blood clots while maintaining hemostasis. Five coagulation serine proteases,factor VII (F7), factor IX (F9), factor X (F10), protein C (PROC), and thrombin (F2),exhibit high sequence similarities and all require vitamin K. All five of these were incorporated into an interactive database of mutations named CoagMDB (http://www.coagMDB.org; last accessed: 9 August 2007). The large number of mutations involved (especially for factor IX) and the increasing problem of out-of-date databases required the development of new database management tools. A text mining tool automatically scans full-length references to identify and extract mutations. High recall rates between 96 and 99% and precision rates of 87 to 93% were achieved. Text mining significantly reduces the time and expertise required to maintain the databases and offers a solution to the problem of locus-specific database management and upkeep. A total of 875 mutations were extracted from 1,279 literature sources. Of these, 116 correspond to Gla domains, 86 to the N-terminal EGF domain, 73 to the C-terminal EGF domain, and 477 to the serine protease domain. The combination of text mining and consensus domain structures enables mutations to be correlated with experimentally-measurable phenotypes based on either low protein levels (Type I) or reduced functional activities (Type II), respectively. A tendency for the conservation of phenotype with structural location was identified. Hum Mutat 29(3), 333,344, 2008. © 2007 Wiley-Liss, Inc. [source]

Comparative analysis of the widespread and conserved PB1-like viruses infecting Pseudomonas aeruginosa

Pieter-Jan Ceyssens
Summary We examined the genetic diversity of lytic Pseudomonas aeruginosa bacteriophage PB1 and four closely related phages (LBL3, LMA2, 14-1 and SN) isolated throughout Europe. They all encapsulate linear, non-permuted genomes between 64 427 and 66 530 bp within a solid, acid-resistant isometric capsid (diameter: 74 nm) and carry non-flexible, contractile tails of approximately 140 nm. The genomes are organized into at least seven transcriptional blocks, alternating on both strands, and encode between 88 (LBL3) and 95 (LMA2) proteins. Their virion particles are composed of at least 22 different proteins, which were identified using mass spectrometry. Post-translational modifications were suggested for two proteins, and a frameshift hotspot was identified within ORF42, encoding a structural protein. Despite large temporal and spatial separations between phage isolations, very high sequence similarity and limited horizontal gene transfer were found between the individual viruses. These PB1-like viruses constitute a new genus of environmentally very widespread phages within the Myoviridae. [source]

Purification of three aminotransferases from Hydrogenobacter thermophilus TK-6 , novel types of alanine or glycine aminotransferase

FEBS JOURNAL, Issue 8 2010
Enzymes, catalysis
Aminotransferases catalyse synthetic and degradative reactions of amino acids, and serve as a key linkage between central carbon and nitrogen metabolism in most organisms. In this study, three aminotransferases (AT1, AT2 and AT3) were purified and characterized from Hydrogenobacter thermophilus, a hydrogen-oxidizing chemolithoautotrophic bacterium, which has been reported to possess unique features in its carbon and nitrogen anabolism. AT1, AT2 and AT3 exhibited glutamate:oxaloacetate aminotransferase, glutamate:pyruvate aminotransferase and alanine:glyoxylate aminotransferase activities, respectively. In addition, both AT1 and AT2 catalysed a glutamate:glyoxylate aminotransferase reaction. Interestingly, phylogenetic analysis showed that AT2 belongs to aminotransferase family IV, whereas known glutamate:pyruvate aminotransferases and glutamate:glyoxylate aminotransferases are members of family I,. In contrast, AT3 was classified into family I, distant from eukaryotic alanine:glyoxylate aminotransferases which belong to family IV. Although Thermococcus litoralis alanine:glyoxylate aminotransferase is the sole known example of family I alanine:glyoxylate aminotransferases, it is indicated that this alanine:glyoxylate aminotransferase and AT3 are derived from distinct lineages within family I, because neither high sequence similarity nor putative substrate-binding residues are shared by these two enzymes. To our knowledge, this study is the first report of the primary structure of bacterial glutamate:glyoxylate aminotransferase and alanine:glyoxylate aminotransferase, and demonstrates the presence of novel types of aminotransferase phylogenetically distinct from known eukaryotic and archaeal isozymes. [source]

The periplasmic peptidyl prolyl cis,trans isomerases PpiD and SurA have partially overlapping substrate specificities

FEBS JOURNAL, Issue 13 2008
Krista H. Stymest
One of the rate-limiting steps in protein folding has been shown to be the cis,trans isomerization of proline residues, catalysed by a range of peptidyl prolyl cis,trans isomerases (PPIases). In the periplasmic space of Escherichia coli and other Gram-negative bacteria, two PPIases, SurA and PpiD, have been identified, which show high sequence similarity to the catalytic domain of the small PPIase parvulin. This observation raises a question regarding the biological significance of two apparently similar enzymes present in the same cellular compartment: do they interact with different substrates or do they catalyse different reactions? The substrate-binding motif of PpiD has not been characterized so far, and no biochemical data were available on how this folding catalyst recognizes and interacts with substrates. To characterize the interaction between model peptides and the periplasmic PPIase PpiD from E. coli, we employed a chemical crosslinking strategy that has been used previously to elucidate the interaction of substrates with SurA. We found that PpiD interacted with a range of model peptides independently of whether they contained proline residues or not. We further demonstrate here that PpiD and SurA interact with similar model peptides, and therefore must have partially overlapping substrate specificities. However, the binding motif of PpiD appears to be less specific than that of SurA, indicating that the two PPIases might interact with different substrates. We therefore propose that, although PpiD and SurA have partially overlapping substrate specificities, they fulfil different functions in the cell. [source]

Nocardiosis in large yellow croaker, Larimichthys crocea (Richardson)

G-L Wang
Abstract An epizootic in seawater-cage reared large yellow croaker, Larimichthys crocea, in China was caused by a Nocardia sp. from August to October 2003. The cumulative mortality rate was 15% and the diseased fish were 16 months old with individual length varying from 25 to 30 cm. Multiple, white nodules, 0.1,0.2 cm in diameter, were scattered on the heart, spleen and kidney. The morphology of isolated bacteria from Lowenstein,Jensen medium and tryptic soy agar was bead-like or long, slender, filamentous rods. Experimental infection indicated that the isolated bacterium was the pathogen responsible for the mortalities. A partial sequence of the 16S rRNA gene of the organism and the type strain of Nocardia seriolae JCM 3360T (Z36925) formed a monophyletic clade with a high sequence similarity of 99.9%. Based on the morphological, physiological, biological properties and the phylogenetic analysis, the pathogenic organism was identified as N. seriolae. This is the first report on N. seriolae -infected large yellow croaker in aquaculture. [source]

Binding of platelet glycoprotein Ib, through the convex surface of leucine-rich repeats domain of glycoprotein IX

Summary.,Background: The mechanism of assembly of the platelet glycoprotein (GP) Ib-IX complex from GPIb,, GPIb, and GPIX subunits is not entirely clear. In this complex, ectodomains of both GPIb, and GPIX subunits contain two leucine-rich repeats (LRR) and share high sequence similarity. However, they differ noticeably in stability, hampering further analysis of their interaction. Objectives and methods: Guided by analysis of the LRR structure, we report a well-folded Ib,/IX chimera and its usage in dissecting GPIX function. Results: In this chimera, three non-contiguous sequences that may constitute the putative convex surface of the GPIb, ectodomain are replaced by their GPIX counterparts. Like GPIb, but unlike GPIX ectodomain, it can secrete from transfected Chinese hamster ovary cells and fold into a stable conformation. Furthermore, replacing the ectodomain in GPIX with the Ib,/IX chimera, but not the GPIb, ectodomain, preserved its interaction with GPIb, as demonstrated by its native-like GPIb,-induced increase in surface expression and coimmunoprecipitation. Conclusions: The putative convex surface of the LRR domain in GPIX is sufficient, in the context of full-length subunit, to mediate its association with GPIb,. [source]

Modelling cross-hybridization on phylogenetic DNA microarrays increases the detection power of closely related species

Abstract DNA microarrays are a popular technique for the detection of microorganisms. Several approaches using specific oligomers targeting one or a few marker genes for each species have been proposed. Data analysis is usually limited to call a species present when its oligomer exceeds a certain intensity threshold. While this strategy works reasonably well for distantly related species, it does not work well for very closely related species: Cross-hybridization of nontarget DNA prevents a simple identification based on signal intensity. The majority of species of the same genus has a sequence similarity of over 90%. For biodiversity studies down to the species level, it is therefore important to increase the detection power of closely related species. We propose a simple, cost-effective and robust approach for biodiversity studies using DNA microarray technology and demonstrate it on scenedesmacean green algae. The internal transcribed spacer 2 (ITS2) rDNA sequence was chosen as marker because it is suitable to distinguish all eukaryotic species even though parts of it are virtually identical in closely related species. We show that by modelling hybridization behaviour with a matrix algebra approach, we are able to identify closely related species that cannot be distinguished with a threshold on signal intensity. Thus this proof-of-concept study shows that by adding a simple and robust data analysis step to the evaluation of DNA microarrays, species detection can be significantly improved for closely related species with a high sequence similarity. [source]

Characterization of an acyl-CoA: carboxylate CoA-transferase from Aspergillus nidulans involved in propionyl-CoA detoxification

Christian B. Fleck
Summary Filamentous fungi metabolize toxic propionyl-CoA via the methylcitrate cycle. Disruption of the methylcitrate synthase gene leads to an accumulation of propionyl-CoA and attenuates virulence of Aspergillus fumigatus. However, addition of acetate, but not ethanol, to propionate-containing medium strongly reduces the accumulation of propionyl-CoA and restores growth of the methylcitrate synthase mutant. Therefore, the existence of a CoA-transferase was postulated, which transfers the CoASH moiety from propionyl-CoA to acetate and, thereby, detoxifying the cell. In this study, we purified the responsible protein from Aspergillus nidulans and characterized its biochemical properties. The enzyme used succinyl-, propionyl- and acetyl-CoA as CoASH donors and the corresponding acids as acceptor molecules. Although the protein displayed high sequence similarity to acetyl-CoA hydrolases this activity was hardly detectable. We additionally identified and deleted the coding DNA sequence of the CoA-transferase. The mutant displayed weak phenotypes in the presence of propionate and behaved like the wild type when no propionate was present. However, when a double-deletion mutant defective in both methylcitrate synthase and CoA-transferase was constructed, the resulting strain was unable to grow on media containing acetate and propionate as sole carbon sources, which confirmed the in vivo activity of the CoA-transferase. [source]

Functional analysis of the Alternaria brassicicola non-ribosomal peptide synthetase gene AbNPS2 reveals a role in conidial cell wall construction

SUMMARY Alternaria brassicicola is a necrotrophic pathogen causing black spot disease on virtually all cultivated Brassica crops worldwide. In many plant pathosystems fungal secondary metabolites derived from non-ribosomal peptide synthetases (NPSs) are phytotoxic virulence factors or are antibiotics thought to be important for niche competition with other micro-organisms. However, many of the functions of NPS genes and their products are largely unknown. In this study, we investigated the function of one of the A. brassicicola NPS genes, AbNPS2. The predicted amino acid sequence of AbNPS2 showed high sequence similarity with A. brassicae, AbrePsy1, Cochliobolus heterostrophus, NPS4 and a Stagonospora nodorum NPS. The AbNPS2 open reading frame was predicted to be 22 kb in length and encodes a large protein (7195 amino acids) showing typical NPS modular organization. Gene expression analysis of AbNPS2 in wild-type fungus indicated that it is expressed almost exclusively in conidia and conidiophores, broadly in the reproductive developmental phase. AbNPS2 gene disruption mutants showed abnormal spore cell wall morphology and a decreased hydrophobicity phenotype. Conidia of abnps2 mutants displayed an aberrantly inflated cell wall and an increase in lipid bodies compared with wild-type. Further phenotypic analyses of abnps2 mutants showed decreased spore germination rates both in vitro and in vivo, and a marked reduction in sporulation in vivo compared with wild-type fungus. Moreover, virulence tests on Brassicas with abnps2 mutants revealed a significant reduction in lesion size compared with wild-type but only when aged spores were used in experiments. Collectively, these results indicate that AbNPS2 plays an important role in development and virulence. [source]

Sequence similarities between Raspberry leaf mottle virus, Raspberry leaf spot virus and the closterovirus Raspberry mottle virus

W.J. McGavin
A sequencing study was performed to determine the relationship between Raspberry mottle virus (RMoV), a newly identified tentative closterovirus found in the United States, and Raspberry leaf mottle virus (RLMV) and Raspberry leaf spot virus (RLSV), which have been known for many years to be components of Raspberry mosaic disease (RMD) in the UK and Europe but which have not been characterised at the molecular level. Cloning and sequencing of cDNAs amplified by reverse transcription-PCR revealed the presence of closteroviruses with high sequence similarity to RMoV in infected plants from the SCRI Rubus virus collection, as well as in a number of samples collected from RMD-symptomatic raspberry plants located at different farms in Scotland and England. These results suggest that RMoV, RLMV and RLSV are isolates of the same virus and we propose that they all should be referred to as RLMV, which was the first of these viruses to be described. Many of the field samples were also infected with a second closterovirus isolate, parts of which could be amplified using RLMV-derived primers. The coat protein amino acid sequences of RLMV and the second virus (PM1) were only 78% identical, even though helicase domain and RNA-dependent RNA polymerase (RDRP) domain sequences were more than 97% identical between RLMV and PM1. [source]