bluuebunny/crossref_metadata_2025 huggingface dataset:
Processing of natural language queries to a relational database
Authors: Samsonova M. , Pisarev A. , Blagov M.
Published: July 2003

Motivation: A new method is developed to query a relational database in natural language (NL).

Results: The method, based on a semantic approach, interprets grammatical and lexical units of a natural language into concepts of subject domain, which are given in a conceptual scheme. The conceptual scheme is mapped formally onto the logical scheme. We applied the method to query the FlyEx database in natural language. FlyEx contains information on the expression of segmentation genes in Drosophila melanogaster. The method allows formulation of queries in various natural languages simultaneously, and is adaptive to changes in the knowledge domain and user’s views. It provides optimal transformation of queries from natural language to SQL, as well as visualization of information as a hyperscheme. The method does not require specification of all possible language constructions as well as a standard grammar accuracy in formulation of NL queries.

Availability: http://urchin.spbcas.ru/NLP/NLP.html

Contact: samson@spbcas.ru

Keywords: natural language processing, relational databases, conceptual scheme, segmentation genes, D.melanogaster, query optimization.

*To whom correspondence should be addressed.
A tool to assist the study of specific
features at protein binding sites
Authors: Santander Víctor , Portales Miguel Angel , Melo Francisco
Published: July 2003

The Protein Data Bank contains a large amount of proteins that have been solved with small ligands bound to them. This constitutes a rich source of information for the study of the specific requirements of protein sites to bind small molecules with a favorable free energy. The specific atomic composition and three-dimensional geometric restraints of protein binding sites for different ligands could be easily obtained from there. The development of accurate binding site descriptors in proteins constitutes a valuable tool to assist in the large-scale prediction and annotation of protein function in whole genomes. In this work, an integrated database containing some processed and calculated protein/ligand information is described. It is expected that this database will constitute a useful tool for people working in the prediction of protein function from its structure. The database is accessible from the Internet through a web server located at: http://protein.bio.puc.cl

Contact: fmelo@genes.bio.puc.cl

Keywords: Ligands, function, structure, proteins, binding sites, conformational analysis

*To whom correspondence should be addressed.
STRUCLA: a WWW meta-server for protein structure comparison and
evolutionary classification
Authors: Sasin Joanna M. , Kurowski Michal A. , Bujnicki Janusz M.
Published: July 2003

Motivation: Evolutionary relationships of proteins have long been derived from the alignment of protein sequences. But from the view of function, most restraints of evolutionary divergence operate at the level of tertiary structure. It has been demonstrated that quantitative measures of dissimilarity in families of structurally similar proteins can be applied to the construction of trees from a comparison of their three-dimensional structures. However, no convenient tool is publicly available to carry out such analyses.

Results: We developed STRUCLA (STRUcture CLAssification), a WWW tool for generation of trees based on evolutionary distances inferred from protein structures according to various methods. The server takes as an input a list of PDB files or the initial alignment of protein coordinates provided by the user (for instance exported from SWISS PDB VIEWER). The user specifies the distance cutoff and selects the distance measures. The server returns series of unrooted trees in the NEXUS format and corresponding distance matrices, as well as a consensus tree. The results can be used as an alternative and a complement to a fixed hierarchy of current protein structure databases. It can complement sequence-based phylogenetic analysis in the ‘twilight zone of homology’, where amino acid sequences are too diverged to provide reliable relationships.

Availability: The service is free for all users and available at http://asia.genesilico.pl/strucla/

Contact: asia@genesilico.pl; iamb@genesilico.pl

*To whom correspondence should be addressed.
Discovering molecular pathways from protein interaction
and gene expression data
Authors: Segal E. , Wang H. , Koller D.
Published: July 2003

In this paper, we describe an approach for identifying‘ pathways’ from gene expression and protein interaction data. Our approach is based on the assumption that many pathways exhibit two properties: their genes exhibit a similar gene expression profile, and the protein products of the genes often interact. Our approach is based on a unified probabilistic model, which is learned from the data using the EM algorithm. We present results on two Saccharomyces cerevisiae gene expression data sets, combined with a binary protein interaction data set. Our results show that our approach is much more successful than other approaches at discovering both coherent functional groups and entire protein complexes.

Contact: eran@cs.stanford.edu

Keywords: probabilistic models, protein interaction, gene expression.

*To whom correspondence should be addressed.
Genome-wide discovery of transcriptional modules from
DNA sequence and gene expression
Authors: Segal E. , Yelensky R. , Koller D.
Published: July 2003

In this paper, we describe an approach for understanding transcriptional regulation from both gene expression and promoter sequence data. We aim to identify transcriptional modules—sets of genes that are co-regulated in a set of experiments, through a common motif profile. Using the EM algorithm, our approach refines both the module assignment and the motif profile so as to best explain the expression data as a function of transcriptional motifs. It also dynamically adds and deletes motifs, as required to provide a genome-wide explanation of the expression data. We evaluate the method on two Saccharomyces cerevisiae gene expression data sets, showing that our approach is better than a standard one at recovering known motifs and at generating biologically coherent modules. We also combine our results with binding localization data to obtain regulatory relationships with known transcription factors, and show that many of the inferred relationships have support in the literature.

Contact: eran@cs.stanford.edu

Keywords: probabilistic models, gene expression, transcriptional regulation.

*To whom correspondence should be addressed.
CREME: a framework for identifying cis-regulatory modules
in human-mouse conserved segments
Authors: Sharan Roded , Ovcharenko Ivan , Ben-Hur Asa , Karp Richard M.
Published: July 2003

Motivation: The binding of transcription factors to specific regulatory sequence elements is a primary mechanism for controlling gene transcription. Recent findings suggest a modular organization of binding sites for transcription factors that cooperate in the regulation of genes. In this work we establish a framework for finding recurrent cis-regulatory modules in the promoters of a selected set of genes and scoring their statistical significance.

Results: Proceeding from a database of identified binding site motifs and their genomic locations we seek motifs whose frequency in the selected promoters is different than in a background promoter set. We present several statistical tests designed for this purpose. We provide a hashing algorithm for detecting combinations of these motifs that co-occur in clusters within the selected promoters. The significance of such co-occurrences is evaluated using novel statistical scores. Our methods are combined in CREME, a suite of software which includes a browser for viewing the pattern of occurrence of selected cis-regulatory modules. We applied our methodology to find modules within human-mouse conserved promoter segments, focusing on cell cycle regulated genes and stress response related genes. To validate the biological significance of the identified modules we tested whether the associated genes tended to be co-expressed or share similar function. In the cell cycle set five of the seven identified sets of genes were coherently expressed. On the stress response data four of the six detected sets fell predominantly into well-defined functional sub-categories.

Availability: http://icsi.berkeley.edu/~roded/creme.html

Contact: roded@icsi.berkeley.edu.

Keywords: Cis-regulatory module, transcription factor binding site, motif cluster, statistical test.

*To whom correspondence should be addressed.

†These authors contributed equally to this work.
A probabilistic method to detect regulatory modules
Authors: Sinha Saurabh , van Nimwegen Erik , Siggia Eric D.
Published: July 2003

Motivation: The discovery of cis-regulatory modules in metazoan genomes is crucial for understanding the connection between genes and organism diversity.

Results: We develop a computational method that uses Hidden Markov Models and an Expectation Maximization algorithm to detect such modules, given the weight matrices of a set of transcription factors known to work together. Two novel features of our probabilistic model are: (i) correlations between binding sites, known to be required for module activity, are exploited, and (ii) phylogenetic comparisons among sequences from multiple species are made to highlight a regulatory module. The novel features are shown to improve detection of modules, in experiments on synthetic as well as biological data.

Availability: The source code for the programs is available upon request from the authors.

Contact: {saurabh,erik,siggia}@lonnrot.rockefeller.edu

Keywords: hidden Markov model, cis-regulatory modules, motif correlations, phylogenetic comparison

*To whom correspondence should be addressed.
myGrid: personalised
bioinformatics on the information grid
Authors: Stevens Robert D. , Robinson Alan J. , Goble Carole A.
Published: July 2003

Motivation: The myGrid project aims to exploit Grid technology, with an emphasis on the Information Grid, and provide middleware layers that make it appropriate for the needs of bioinformatics. myGrid is building high level services for data and application integration such as resource discovery, workflow enactment and distributed query processing. Additional services are provided to support the scientific method and best practice found at the bench but often neglected at the workstation, notably provenance management, change notification and personalisation.

Results: We give an overview of these services and their metadata. In particular, semantically rich metadata expressed using ontologies necessary to discover, select and compose services into dynamic workflows.

Availability: Software is available on request from the authors and information from http://www.mygrid.org.uk

Contact: carole@cs.man.ac.uk

*To whom correspondence should be addressed.
Scaling up accurate phylogenetic reconstruction from gene-order data
Authors: Tang Jijun , Moret Bernard M.E.
Published: July 2003

Motivation: Phylogenetic reconstruction from gene-order data has attracted increasing attention from both biologists and computer scientists over the last few years. Methods used in reconstruction include distance-based methods (such as neighbor-joining), parsimony methods using sequence-based encodings, Bayesian approaches, and direct optimization. The latter, pioneered by Sankoff and extended by us with the software suite GRAPPA, is the most accurate approach, but cannot handle more than about 15 genomes of limited size (e.g. organelles).

Results: We report here on our successful efforts to scale up direct optimization through a two-step approach: the first step decomposes the dataset into smaller pieces and runs the direct optimization (GRAPPA) on the smaller pieces, while the second step builds a tree from the results obtained on the smaller pieces. We used the sophisticated disk-covering method (DCM) pioneered by Warnow and her group, suitably modified to take into account the computational limitations of GRAPPA. We find that DCM-GRAPPA scales gracefully to at least 1000 genomes of a few hundred genes each and retains surprisingly high accuracy throughout the range: in our experiments, the topological error rate rarely exceeded a few percent. Thus, reconstruction based on gene-order data can now be accomplished with high accuracy on datasets of significant size.

Availability: All of our software is available in source form under GPL at http://www.compbio.unm.edu

Contact: moret@cs.unm.edu

*To whom correspondence should be addressed.
Skew in CG content near the transcription start site in
Arabidopsis thaliana
Authors: Tatarinova Tatiana , Brover Vyacheslav , Troukhan Maxim , Alexandrov Nickolai
Published: July 2003

We have discovered a novel statistical feature of Arabidopsis thaliana genome that remarkably correlates with a position of transcription start site—CG skew peak. We hypothesize that the phenomenon can be explained by the higher mutability of unprotected cytosines.

Keywords: CG skew, promoter, transcription, mutation, full-length cDNA

Contact: nicka@ceres-inc.com

*To whom correspondence should be addressed.
Extraction of correlated gene clusters from multiple genomic data
by generalized kernel canonical correlation analysis
Authors: Yamanishi Y. , Vert J.-P. , Nakaya A. , Kanehisa M.
Published: July 2003

Motivation: A major issue in computational biology is the reconstruction of pathways from several genomic datasets, such as expression data, protein interaction data and phylogenetic profiles. As a first step toward this goal, it is important to investigate the amount of correlation which exists between these data.

Results: These methods are successfully tested on their ability to recognize operons in the Escherichia coli genome, from the comparison of three datasets corresponding to functional relationships between genes in metabolic pathways, geometrical relationships along the chromosome, and co-expression relationships as observed by gene expression data.

Contact: yoshi@kuicr.kyoto-u.ac.jp

*To whom correspondence should be addressed.
Evaluation of text data mining for database curation:
lessons learned from the KDD Challenge Cup
Authors: Yeh Alexander S. , Hirschman Lynette , Morgan Alexander A.
Published: July 2003

Motivation: The biological literature is a major repository of knowledge. Many biological databases draw much of their content from a careful curation of this literature. However, as the volume of literature increases, the burden of curation increases. Text mining may provide useful tools to assist in the curation process. To date, the lack of standards has made it impossible to determine whether text mining techniques are sufficiently mature to be useful.

Results: We report on a Challenge Evaluation task that we created for the Knowledge Discovery and Data Mining (KDD) Challenge Cup. We provided a training corpus of 862 articles consisting of journal articles curated in FlyBase, along with the associated lists of genes and gene products, as well as the relevant data fields from FlyBase. For the test, we provided a corpus of 213 new (‘blind’) articles; the 18 participating groups provided systems that flagged articles for curation, based on whether the article contained experimental evidence for gene expression products. We report on the evaluation results and describe the techniques used by the top performing groups.

Contact: asy@mitre.org

Keywords: text mining, evaluation, curation, genomics, data management

*To whom correspondence should be addressed.
Extracting synonymous gene and protein terms from biological literature
Authors: Yu Hong , Agichtein Eugene
Published: July 2003

Motivation: Genes and proteins are often associated with multiple names. More names are added as new functional or structural information is discovered. Because authors can use any one of the known names for a gene or protein, information retrieval and extraction would benefit from identifying the gene and protein terms that are synonyms of the same substance.

Results: We have explored four complementary approaches for extracting gene and protein synonyms from text, namely the unsupervised, partially supervised, and supervised machine-learning techniques, as well as the manual knowledge-based approach. We report results of a large scale evaluation of these alternatives over an archive of biological journal articles. Our evaluation shows that our extraction techniques could be a valuable supplement to resources such as SWISSPROT, as our systems were able to capture gene and protein synonyms not listed in the SWISSPROT database.

Data Availability: The extracted gene and protein synonyms are available at http://synonyms.cs.columbia.edu/

Contact: {hongyu,eugene}@cs.columbia.edu

*To whom correspondence should be addressed.
Identification of genes in the tomato big bud phytoplasma and comparison to those in sweet potato little leaf-V4 phytoplasma
Authors: Streten Claire , Gibb Karen S.
Published: July 2003

Genetic relatedness of phytoplasmas is commonly defined on the basis of differences in the highly conserved 16S rRNA gene, which may not resolve closely related phytoplasmas. An example of this is the closely related tomato big bud (TBB) and sweet potato little leaf strain V4 (SPLL-V4) phytoplasmas, which cannot easily be differentiated by their 16S rRNA gene sequences. This study aimed to identify genes on the TBB phytoplasma chromosome which could be used to examine genetic variation between these two closely related phytoplasmas. Random clones generated from TBB phytoplasma genomic DNA were sequenced and characterized by database analysis. Twenty-three genes were identified within 19 random clones, which contained approximately 18·0 kbp of TBB phytoplasma genomic DNA. Half of the TBB phytoplasma genes identified were involved in DNA replication, transcription and translation. The remaining TBB phytoplasma genes were involved in protein secretion, cellular processes and energy metabolism. Phylogenetic analysis of representative genes showed that the TBB phytoplasma grouped with the mycoplasmas with the exception of the TBB phytoplasma secA gene, which grouped with the onion yellows phytoplasma. PCR primers were designed based on the new genes and tested on isolates of the TBB and SPLL-V4 phytoplasmas. Most primers amplified a product from TBB and SPLL-V4 phytoplasma samples. When amplified products were subjected to RFLP analysis, the restriction patterns were the same as the respective original clones. This result confirmed that the same sequence had been amplified by PCR and showed that these isolates were indistinguishable using the new genes. This study showed that in fact the TBB and SPLL-V4 phytoplasmas are closely related even with the analysis of new genes. These new genes have, however, provided insight into the biology of the TBB and SPLL-V4 phytoplasmas.
Haemagglutinin/protease expression and mucin gel penetration in El Tor biotype Vibrio cholerae
Authors: Silva Anisia J. , Pham Kim , Benitez Jorge A.
Published: July 2003

Vibrio cholerae of both biotypes produce a soluble Zn2+-dependent metalloprotease: haemagglutinin/protease (Hap), encoded by hapA. Hap has been shown to have mucinolytic and cytotoxic activity. These activities are likely to play an important role in the pathogenesis of cholera and the reactogenicity of attenuated vaccine strains. Production of Hap requires transcriptional activation by the HapR regulator and is repressed by glucose. The present study shows that mucin purified from two sources, bile salts, and growth at 37 °C enhanced Hap protease production. Analysis of hapA and hapR promoter fusions with the lacZ gene showed both promoters to be activated in a cell-density-dependent pattern. Glucose repressed and mucin induced the hapA promoter by a HapR-independent mechanism. Bile had no effect on either hapR or hapA promoter activity. Expression of hapA was required for vibrios to translocate through a mucin-containing gel. These results suggest Hap to play an important role in cholera pathogenesis by promoting mucin gel penetration, detachment and spreading of infection along the gastrointestinal tract.
Posttranslational processing of the xylanase Xys1L from Streptomyces halstedii JM8 is carried out by secreted serine proteases
Authors: Fernández-Abalos José M. , Reviejo Verónica , Díaz Margarita , Rodríguez Sonia , Leal Fernando , Santamaría Ramón I.
Published: July 2003

The xylanase Xys1L fromStreptomyces halstediiJM8 is known to be processed extracellularly, to produce a protein of 33·7 kDa, Xys1S, that retains catalytic activity but not its cellulose-binding capacity. This paper demonstrates that at least five serine proteases isolated fromStreptomycesspp. have the ability to process the xylanase Xys1L. The genes of two of these extracellular serine proteases, denominated SpB and SpC, were cloned fromStreptomyces lividans66 (a strain commonly used as a host for protein secretion), sequenced, and overexpressed inS. lividans; both purified proteases were able to process Xys1Lin vitro. Three other previously reported purifiedStreptomycesserine proteases, SAM-P20, SAM-P26 and SAM-P45, also processed Xys1Lin vitro. The involvement of serine proteases in xylanase processing-degradationin vivowas demonstrated by co-expression of the xylanase gene (xysA) and the gene encoding the serine protease inhibitor (SLPI) fromS. lividans. Co-expression prevented processing and degradation of Xys1L and resulted in a threefold increase in the xylanase activity present in the culture supernatant. SpB and SpC also have the capacity to process other secreted proteins such as p40, a cellulose-binding protein fromS. halstediiJM8, but do not have any clear effect on other secreted proteins such as amylase (Amy) fromStreptomyces griseusand xylanase Xyl30 fromStreptomyces avermitilis.
Patchiness of murein insertion into the sidewall of Escherichia coli
Authors: De Pedro Miguel A. , Schwarz Heinz , Koch Arthur L.
Published: July 2003

This paper extends, with computer techniques, the authors' previous work on the kinetics of pole wall and sidewall synthesis in Escherichia coli. These findings extend the conclusion that the nascent poles are made of entirely new material and that no new material is inserted into old poles. This requires re-evaluation of ideas in the literature about wall growth and cell division. Mechanisms of various types have been suggested for the growth of Gram-negative rod-shaped bacteria and these will also require major re-evaluation because of the finding, reported here, that the sidewall is made in several modes: patches of new murein, bands of new material largely going circumferentially around the cell, and areas of the sidewall that are enlarged by an intimate and regular admixture of new with the old muropeptides.
Helicobacter pylori tissue tropism: mouse-colonizing strains can target different gastric niches
Authors: Akada Junko K. , Ogura Keiji , Dailidiene Daiva , Dailide Giedrius , Cheverud James M. , Berg Douglas E.
Published: July 2003

Studies with the mouse-adapted Helicobacter pylori strain SS1 had supported an idea that infections by this pathogen start in the gastric antrum and spread to the corpus after extensive mucosal damage. This paper shows that the unrelated strain X47 colonizes the corpus preferentially. Differences between strains in preferred gastric region were detected by co-inoculating mice with a mixture of SS1 and X47, and genotyping H. pylori recovered after 2–8 weeks of infection by vacA s allele PCR and RAPD fingerprinting. Mixed infections were found in each of 59 co-inoculated young C57BL/6J mice. On average, however, SS1 was fourfold more abundant than X47 in the antrum and X47 was threefold more abundant than SS1 in the corpus. Similar results were obtained in mice inoculated first with one strain and then the other strain 2 weeks later. SS1 was even more abundant in the antrum of elderly (>1 year old) mice (97 % of isolates). Qualitatively similar SS1 and X47 tissue distributions were seen using unrelated mouse lines (AKR/J, A/J, DBA/2J, BALB/cJ, LG/J, SM/J), but with significantly different SS1 : X47 ratios in some cases. These results suggest the existence of at least two distinct gastric niches whose characteristics may be affected by host genotype and age (physiology), and indicate that strains differ in how effectively they colonize each niche. Differences among gastric regions and the mixed infections that these allow may contribute to H. pylori diversity and genome evolution.
Antigenic diversity of meningococcal enterobactin receptor FetA, a vaccine component
Authors: Thompson Emily A. L. , Feavers Ian M. , Maiden Martin C. J.
Published: July 2003

Meningococcal FetA (FrpB), an iron-regulated outer-membrane protein and vaccine component, was shown to be highly diverse: a total of 60fetAalleles, encoding 56 protein sequences, were identified from 107 representativeNeisseria meningitidisisolates. Phylogenetic analysis established that the allelic variants had been generated by both point mutation and horizontal genetic exchange. Nucleotide substitution was unevenly distributed in the gene, which contained both conserved and variable sequence regions. The most conserved region of the translated peptide sequence corresponded to an amino-terminal domain of the protein and the most diverse region to a previously identified variable region (VR). A nomenclature system for the peptides encoded by the VR was devised which classified 24 variants into 5 FetA variant families. On the basis of these data, murine polyclonal sera specific for four FetA variants were generated. The reactivities of these sera in whole-cell ELISA experiments were consistent with the hypothesis that the VR encoded an immunodominant epitope and indicated that the sera reacted mainly with variants against which they were raised. The diversity of this protein is likely to limit its effectiveness as a vaccine component.
Transcriptional, translational and metabolic regulation of glycolysis in Lactococcus lactis subsp. cremoris MG 1363 grown in continuous acidic cultures
Authors: Even Sergine , Lindley Nic D. , Cocaign-Bousquet Muriel
Published: July 2003

The physiological behaviour ofLactococcus lactissubsp.cremorisMG 1363 was characterized in continuous culture under various acidic conditions (pH 4·7–6·6). Biomass yield was diminished in cultures with low pH and the energy dedicated to maintenance increased due to organic acid inhibition and cytoplasmic acidification. Under such acidic conditions, the specific rate of glucose consumption by the bacterium increased, thereby enhancing energy supply. This acceleration of glycolysis was regulated by both an increase in the concentrations of glycolytic enzymes (hierarchical regulation) and the specific modulation of enzyme activities (metabolic regulation). However, when the inhibitory effect of intracellular pH on enzyme activity was taken into account in the model of regulation, metabolite regulation was shown to be the dominant factor controlling pathway flux. The changes in glycolytic enzyme concentrations were not correlated directly to modifications in transcript concentrations. Analyses of the relative contribution of the phenomena controlling enzyme synthesis indicated that translational regulation had a major influence compared to transcriptional regulation. An increase in the translation efficiency was accompanied by an important decrease of total cellular RNA concentrations, confirming that the translation apparatus ofL. lactiswas optimized under acid stress conditions.
An enhanced GFP reporter system to monitor gene expression in Borrelia burgdorferi
Authors: Carroll James A. , Stewart Philip E. , Rosa Patricia , Elias Abdallah F. , Garon Claude F.
Published: July 2003

Borrelia burgdorferi regulates genes in response to a number of environmental signals such as temperature and pH. A green fluorescent protein (GFP) reporter system using the ospC, ospA and flaB promoters from B. burgdorferi B31 was introduced into infectious clonal isolates of strains B31 and N40 to monitor and compare gene expression in response to pH and temperature in vitro. GFP could be assayed by epifluorescence microscopy, immunoblotting or spectrofluorometry and was an accurate reporter of target gene expression. It was determined that only 179 bp 5′ of ospC was sufficient to regulate the reporter gfp
in vitro in response to pH and temperature in B. burgdorferi B31. The loss of linear plasmid (lp) 25, lp28-1, lp36 and lp56 had no effect on the ability of B. burgdorferi B31 to regulate ospC in response to pH or temperature. The amount of OspC in N40 transformants was unaffected by changes in pH or temperature of the culture medium. This suggests that regulation of gene expression in response to pH and temperature may vary between these two B. burgdorferi strains.
Production of the signalling molecule, autoinducer-2, by Neisseria meningitidis: lack of evidence for a concerted transcriptional response
Authors: Dove Joanne E. , Yasukawa Kazutoyo , Tinsley Colin R. , Nassif Xavier
Published: July 2003

Neisseria meningitidis is a Gram-negative bacterium which is an important causative agent of septicaemia and meningitis. LuxS has been shown to be involved in the biosynthesis of a quorum sensing molecule, autoinducer-2 (AI-2), known to play a role in virulence in Escherichia coli, as well as other bacteria. Evidence that serogroup B of N. meningitidis produces AI-2, along with the observation that a luxS mutant of this strain had attenuated virulence in an infant rat model of bacteraemia, led to further investigation of the role of this quorum sensing molecule in N. meningitidis. In this study, it is demonstrated that AI-2 is not involved in regulating growth of meningococci, either in culture or in contact with epithelial cells. Furthermore, transcriptional profiling using DNA microarrays shows an absence of the concerted regulation seen in other bacteria. Taken together, these data suggest that in N. meningitidis, AI-2 may be a metabolic by-product and not a cell-to-cell signalling molecule.
Phospholipid composition of several clinically relevant Corynebacterium species as determined by mass spectrometry: an unusual fatty acyl moiety is present in inositol-containing phospholipids of Corynebacterium urealyticum
Authors: Yagüe Genoveva , Segovia Manuel , Valero-Guillén Pedro L.
Published: July 2003

A comparative study on phospholipids ofCorynebacterium amycolatum,Corynebacterium jeikeiumandCorynebacterium urealyticumwas carried out using fast-atom bombardment (FAB) and electrospray ionization (ESI) mass spectrometry. Data obtained indicate the presence of acylphosphatidylglycerol (APG), diphosphatidylglycerol, phosphatidylglycerol (PG), phosphatidylinositol (PI) and triacylphosphatidylinositol dimannosides (Ac3PIM2) in these bacteria. In general, octadecenoyl and hexadecanoyl fatty acyl moieties predominated in phospholipids ofC. amycolatum, whereas high levels of hexadecenoyl were found inC. jeikeiumandC. urealyticum. Mass spectra from purified APG and PG indicated that thesn-1 position of the glycerol was occupied by octadecenoyl in the three species studied. Notably, several major molecular species of PI and Ac3PIM2fromC. urealyticumcontained significant amounts of a moiety identified as 10-methyleneoctadecanoyl, located at thesn-1 position of these molecules. On the other hand, multiantibiotic resistant and susceptible strains ofC. amycolatumdiffered in several minor phospholipid fatty acids of 19 carbon atoms, identified as 10-methyloctadecenoic, 10-methyloctadecanoic (tuberculostearic acid) and 10-methyleneoctadecanoic. The results demonstrate an overall similarity among the phospholipids of the different species studied but also significant differences related to the acyl chains of the glycerol moiety of these compounds, notably the high levels of an unusual fatty acyl moiety in inositol-containing phospholipids ofC. urealyticum.
The sulfane sulfur of persulfides is the actual substrate of the sulfur-oxidizing enzymes from Acidithiobacillus and Acidiphilium spp.
Authors: Rohwerder Thore , Sand Wolfgang
Published: July 2003

To identify the actual substrate of the glutathione-dependent sulfur dioxygenase (EC 1.13.11.18) elemental sulfur oxidation of the meso-acidophilicAcidithiobacillus thiooxidansstrains DSM 504 and K6,Acidithiobacillus ferrooxidansstrain R1 andAcidiphilium acidophilumDSM 700 was analysed. Extraordinarily high specific sulfur dioxygenase activities up to 460 nmol min−1(mg protein)−1were found in crude extracts. All cell-free systems oxidized elemental sulfur only via glutathione persulfide (GSSH), a non-enzymic reaction product from glutathione (GSH) and elemental sulfur. Thus, GSH plays a catalytic role in elemental sulfur activation, but is not consumed during enzymic sulfane sulfur oxidation. Sulfite is the first product of sulfur dioxygenase activity; it further reacted non-enzymically to sulfate, thiosulfate or glutathioneS-sulfonate (). Free sulfide was not oxidized by the sulfur dioxygenase. Persulfide as sulfur donor could not be replaced by other sulfane-sulfur-containing compounds (thiosulfate, polythionates, bisorganyl-polysulfanes or monoarylthiosulfonates). The oxidation of H2S by the dioxygenase required GSSG, i.e. the disulfide of GSH, which reacted non-enzymically with sulfide to give GSSH prior to enzymic oxidation. On the basis of these results and previous findings a biochemical model for elemental sulfur and sulfide oxidation inAcidithiobacillusandAcidiphiliumspp. is proposed.
pckA-deficient Mycobacterium bovis BCG shows attenuated virulence in mice and in macrophages
Authors: Liu Keyi , Yu Jinzhi , Russell David G.
Published: July 2003

Phosphoenolpyruvate carboxykinase (PEPCK) catalyses the reversible decarboxylation and phosphorylation of oxaloacetate (OAA) to form phosphoenolpyruvate (PEP). In this study, the regulation of the PEPCK-encoding genepckAwas examined through the evaluation of green fluorescent protein expression driven by thepckApromoter. The results showed thatpckAwas upregulated by acetate or palmitate but downregulated by glucose. Deletion of thepckAgene ofMycobacteriumbovisBCG led to a reduction in the capacity of the bacteria to infect and survive in macrophages. Moreover, mice infected with ΔpckABCG were able to reduce the bacterial load much more effectively than mice infected with the parental wild-type bacteria. This attenuated virulence was reflected in the degree of pathology, where granuloma formation was diminished both in numbers and degree. The data indicate that PEPCK activity is important during establishment of infection. Whether its role is in the gluconeogenic pathway for carbohydrate formation or in the conversion of PEP to OAA to maintain the TCA cycle remains to be determined.
The haem pigment of the oral anaerobes Prevotella nigrescens and Prevotella intermedia is composed of iron(III) protoporphyrin IX in the monomeric form
Authors: Smalley John W. , Silver Jack , Birss Andrew J. , Withnall Robert , Titler Philip J.
Published: July 2003

The haem pigment of Porphyromonas gingivalis is composed of μ-oxo bishaem, [Fe(III)PPIX]2O, but the nature of that generated by Prevotella species has not been established. Mössbauer, Raman and UV-visible spectrophotometry were used to characterize the haem pigment of Prevotella intermedia and Prevotella nigrescens. Mössbauer and Raman spectroscopy revealed the major haem species to be monomeric iron protoporphyrin IX, Fe(III)PPIX.OH (haematin). The terminal growth pH of both species on blood agar was between 5·8 and 6·0, which favours the formation and maintenance of monomeric Fe(III)PPIX.OH. Incubation of Pr. nigrescens and Pr. intermedia with oxyhaemoglobin at pH 6·5 resulted in formation of aquomethaemoglobin which was degraded to generate Fe(III)PPIX.OH which in turn became cell-associated, whilst incubation at pH 7·5 resulted in formation of [Fe(III)PPIX]2O. It is concluded that both Prevotella species degrade oxyhaemoglobin to form [Fe(III)PPIX]2O as an intermediate, which is converted to Fe(III)PPIX.OH through a depression in pH. The low pH encourages cell-surface deposition of insoluble Fe(III)PPIX.OH which would act as a barrier against oxygen and reactive oxygen species, and also protect against H2O2 through its inherent catalase activity.
The Escherichia coli AIDA autotransporter adhesin recognizes an integral membrane glycoprotein as receptor
Authors: Laarmann Sven , Schmidt M. Alexander
Published: July 2003

The AIDA-I autotransporter adhesin, as a prototype of the AIDA adhesin family, represents a tripartite antigen consisting of the functional adhesin AIDA-I (α-domain), which mediates the specific attachment of bacteria to target cells, and a two-domain translocator (AIDAc) organized in the β
1- and β
2-domains. Cellular receptor moieties for the adhesin AIDA-I have not been identified. Here, it is demonstrated that the purified adhesin binds specifically to a high-affinity class of receptors on HeLa cells. Additionally, the adhesin was found to bind to a variety of mammalian cell types, indicating a broad tissue distribution of the receptor moiety. By using complementary techniques, including co-immunoprecipitation and one- and two-dimensional gel electrophoresis, the AIDA-I binding protein on HeLa cells was identified as a surface glycoprotein of about 119 kDa (gp119). The gp119 AIDA-I cellular receptor protein was characterized biochemically and found to be an integral N-glycosylated membrane protein with a pI of 5·2.
Virulence attenuation of two Mas-like polyketide synthase mutants of Mycobacterium tuberculosis
Authors: Rousseau Cécile , Sirakova Tatiana D. , Dubey Vinod S. , Bordat Yann , Kolattukudy Pappachan E. , Gicquel Brigitte , Jackson Mary
Published: July 2003

The cell envelope of pathogenic mycobacteria is highly distinctive in that it contains a large number of structurally related very long multiple methyl-branched fatty acids. These complex molecules are thought to play important roles in cell envelope organization and virulence. The genetic and enzymic characterization of the polyketide synthase Mas, which is responsible for the synthesis of one such family of fatty acids (the mycocerosic acids), paved the way towards the identification of other enzymes involved in the synthesis of methyl-branched fatty acids in M. tuberculosis. In an effort to elucidate the origin of these complex fatty acids and their possible involvement in pathogenesis, the two mas-like polyketide genes pks5 and pks7 were disrupted in M. tuberculosis and the effects of their inactivation on fatty acid composition and virulence were analysed. While the disruption of pks7 resulted in a mutant deficient in the production of phthiocerol dimycocerosates, the cell envelope composition of the pks5 mutant was found to be identical to that of the wild-type parental strain M. tuberculosis H37Rv. Interestingly, both the pks5 and pks7 mutants displayed severe growth defects in mice.
Involvement of the cgtA gene function in stimulation of DNA repair in Escherichia coli and Vibrio harveyi
Authors: Zielke Ryszard , Sikora Aleksandra , Dutkiewicz Rafał , Wegrzyn Grzegorz , Czyż Agata
Published: July 2003

CgtA is a member of the Obg/Gtp1 subfamily of small GTP-binding proteins. CgtA homologues have been found in various prokaryotic and eukaryotic organisms, ranging from bacteria to humans. Nevertheless, despite the fact thatcgtAis an essential gene in most bacterial species, its function in the regulation of cellular processes is largely unknown. Here it has been demonstrated that in two bacterial species,Escherichia coliandVibrio harveyi, thecgtAgene product enhances survival of cells after UV irradiation. Expression of thecgtAgene was found to be enhanced after UV irradiation of bothE. coliandV. harveyi. Moderate overexpression ofcgtAresulted in higher UV resistance ofE. coliwild-type anddnaQstrains, but not inuvrA,uvrB,umuCandrecAmutant hosts. Overexpression of theE. coli recAgene in theV. harveyi cgtAmutant, which is very sensitive to UV light, restored the level of survival of UV-irradiated cells to the levels observed for wild-type bacteria. Moreover, the basal level of the RecA protein was lower in a temperature-sensitivecgtAmutant ofE. colithan in thecgtA+strain, and contrary to wild-type bacteria, no significant increase inrecAgene expression was observed after UV irradiation of thiscgtAmutant. Finally, stimulation ofuvrBgene transcription under these conditions was impaired in theV. harveyi cgtAmutant. All these results strongly suggest that thecgtAgene product is involved in DNA repair processes, most probably by stimulation ofrecAgene expression and resultant activation of RecA-dependent DNA repair pathways.
Cloning, sequencing and heterologous expression of the medermycin biosynthetic gene cluster of Streptomyces sp. AM-7161: towards comparative analysis of the benzoisochromanequinone gene clusters
Authors: Ichinose Koji , Ozawa Makoto , Itou Keiko , Kunieda Kanako , Ebizuka Yutaka
Published: July 2003

Medermycin is aStreptomycesaromaticC-glycoside antibiotic classified in the benzoisochromanequinones (BIQs), which presents several interesting biosynthetic problems concerning polyketide synthase (PKS), post-PKS tailoring and deoxysugar pathways. The biosynthetic gene cluster for medermycin (themedcluster) was cloned fromStreptomycessp. AM-7161. Completeness of the clone was proved by the heterologous expression of a cosmid carrying the entiremedcluster inStreptomyces coelicolorCH999 to produce medermycin. The DNA sequence of the cosmid (36 202 bp) revealed 34 complete ORFs, with an incomplete ORF at either end. Functional assignment of the deduced products was made for PKS and biosynthetically related enzymes, tailoring steps including strereochemical control, oxidation, angolosamine pathway,C-glycosylation, and regulation. Themedcluster was estimated to be about 30 kb long, covering 29 ORFs. An unusual characteristic of the cluster is the disconnected organization of the minimal PKS genes:med-ORF23 encoding the acyl carrier protein is 20 kb apart frommed-ORF1 andmed-ORF2 for the two ketosynthase components. Secondly, the six genes (med-ORF14, 15, 16, 17, 18 and 20) for the biosynthesis of the deoxysugar, angolosamine, are all contiguous. Finally, the finding of a glycosyltransferase gene,med-ORF8, suggests a possible involvement of conventionalC-glycosylation in medermycin biosynthesis. Comparison among the three complete BIQ gene clusters –medand those for actinorhodin (act) and granaticin (gra) – revealed some common genes whose deduced functions are unavailable from database searches (the ‘unknowns’). An example ismed-ORF5, a homologue ofactVI-ORF3 andgra-ORF18, which was highlighted by a recent proteomic analysis ofS. coelicolorA3(2).
Signal-mediated cross-talk regulates stress adaptation in Vibrio species
Authors: McDougald Diane , Srinivasan Sujatha , Rice Scott A. , Kjelleberg Staffan
Published: July 2003

Quorum sensing systems serve as a means of ‘census taking’ of conspecific and non-conspecific bacteria in the near vicinity. The acylated homoserine lactone (AHL) quorum sensing system has been proposed to be primarily an intra-specific communication system, while the AI-2 autoinducer signalling system is proposed to be an interspecific communication system. Here it is shown that AI-2-like signalling in two marineVibriospecies,Vibrio vulnificusand ‘Vibrio angustum’ S14, induces the core response phenotypes of starvation adaptation and stress resistance, and that a signal antagonist can competitively inhibit these phenotypes. Furthermore, the signals produced by a range ofVibriospecies have the ability to induce these phenotypes inV. vulnificusand ‘V. angustum’ S14, indicating that, at least inVibriospecies, AI-2-like signalling systems function as interspecies communication systems capable of ‘cross-talk’ and of regulating environmentally relevant phenotypes.
Budding of melanized Cryptococcus neoformans in the presence or absence of l-dopa
Authors: Nosanchuk Joshua D. , Casadevall Arturo
Published: July 2003

Cryptococcus neoformansis a pathogenic fungus that produces melanin when incubated in the presence of certain phenolic substrates such asl-3,4-dihydroxyphenylalanine (l-dopa). Melanin is an enigmatic polymer that is deposited in the cell wall and contributes to virulence. Substantial progress has been made in understanding the synthesis of melanin and the mechanisms by which it contributes to virulence, but relatively little is known about how melanin is rearranged during growth and budding. In this study we used transmission and scanning electron microscopy and immunofluorescence of melanized cells and melanin ‘ghosts' to study the process of melanization during replication. Budding in melanizedC. neoformansresults in focal disruption of cell-wall melanin at the bud site. In the presence ofl-dopa, bud-related melanin defects are repaired and daughter cells are melanized. However, in the absence of substrate, mother cells cannot repair their melanin defects and daughter cells are non-melanized. Hence, melanin in the parent cell is not carried to the daughter cells, but rather is synthesizedde novoin buds. These results imply that melanin remodelling occurs during cell growth in a process that involves degradation and synthesis at sites of budding.
Metal-responsive elements in Pleurotus ostreatus laccase gene promoters
Authors: Faraco Vincenza , Giardina Paola , Sannia Giovanni
Published: August 2003

Fungal laccase gene transcription is strongly induced by copper ions; notably, some laccase promoters contain multiple putative metal-responsive elements (MREs). Previously, it has been demonstrated that thePleurotus ostreatuslaccase genespoxcandpoxa1bare transcriptionally induced by copper, and several putative MREs were found in the promoter regions of these genes, which extend for about 400 nt upstream of the start codon (ATG). Identification of MRE sequences, which are protected by protein binding in thepoxcandpoxa1bpromoter regions, has been achieved by footprinting analyses. Electromobility shift assays led to the evaluation of the ability of the identified MREs to bind protein(s), and the role of specific nucleotides of these elements in complex formation has also been analysed. The formation of complexes between analysed MREs and fungal proteins requires the absence of metal ions. Proteins extracted from fungus grown in copper-depleted medium are able to form complexes with MREs, whilst proteins extracted from fungus grown in copper-containing medium are able to form complexes only in the presence of a metal chelator. Moreover, copper-depleted proteins are unable to form complexes when copper or zinc ions are added. UV-cross-linking analyses led to the determination of the molecular masses of the MRE-binding proteins. In thepoxa1bpromoter, a GC-rich region, homologous to the core binding site for transcription factor Sp1, decreases the binding affinity of the adjacent MRE, affecting its interactions with fungal protein factors.
Output and Input Biases Caused by Public Agricultural Research and Extension in Japan, 1957–1997
Authors: Kuroda Yoshimi , Lee Yong‐Sun
Published: July 2003

The present paper investigates the magnitude and the Hicksian output and input biases of technological change brought about by investments in public agricultural research and extension activities in Japan. Given this objective, it estimates a translog multiproduct cost function for 1957–1997. Empirical results show that the cost reduction effects were fairly large. The Hicksian (1932) biases were found to be: (i) livestock‐augmenting; (ii) labor‐ and other‐inputs‐saving; and (iii) machinery‐ and intermediate‐inputs using. Except for other inputs, the directions of the biases are consistent with the Hicksian (1932) induced innovation hypothesis, which supports the public‐sector‐induced‐innovation model proposed by Hayami and Ruttan (1985) and Ohtsuka (1982).