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PubMed2004

Public web-based services from the European Bioinformatics Institute.

The mission of the European Bioinformatics Institute (EBI), an outstation of the European Molecular Biology Laboratory (EMBL) in Heidelberg, is to ensure that the growing body of information from molecular biology and genome research is placed in the public domain and is accessible freely to all parts of the scientific community in ways that promote scientific progress. To fulfil this mission, the EBI provides a wide variety of free, publicly available bioinformatics services. These can be divided into data submissions processing; access to query, analysis and retrieval systems and tools; ftp downloads of software and databases; training and education and user support. All of these services are available at the EBI website: http://www.ebi.ac.uk/services. This paper provides a detailed introduction to the interactive analysis systems that are available from the EBI and a brief introduction to other, related services.

Computational Biology↗
PubMed2006

Genomic analysis of host-pathogen interaction between Fusarium graminearum and wheat during early stages of disease development.

Fusarium graminearum strains responsible for causing the plant disease Fusarium head blight vary greatly in their ability to cause disease and produce mycotoxins on wheat. With the goal of understanding fungal gene expression related to pathogenicity, three cDNA libraries were created by suppression subtractive hybridization using wheat heads inoculated with a highly aggressive strain and either water or a less aggressive strain of this pathogen. Eighty-four fungal genes expressed during initial disease development were identified. The probable functions of 49 of these genes could be inferred by bioinformatic analysis. Thirty-five ESTs had no known homologues in current databases and were not identified by ab initio gene prediction methods. These ESTs from infected wheat heads probably represent F. graminearum genes that previously were not annotated. Four genes represented in one of these libraries were selected for targeted gene replacement, leading to the characterization of a two-component response regulator homologue involved in pathogenicity of the fungus. The mutants for this gene showed reduced sporulation and delayed spread of Fusarium head blight on wheat.

Computational Biology↗
PubMed2006

A survey of protein-protein complex crystallizations.

A survey of crystallization conditions was carried out for 650 published protein-protein complexes in the Protein Data Bank (PDB) of the Research Collaboratory for Structural Bioinformatics (RCSB). This resulted in the establishment of a Protein Complex Crystallization Database (PCCD) and a set of configuration-space boundaries for protein-complex crystallizations. Overall, polyethylene glycol (PEG) based conditions accounted for 70-80% of all crystallizations, with PEG 3000-4000, 5000-6000 and 8000 being the most frequently used. The median values of PEG concentrations were between 10 and 20% and were inversely correlated with their molecular weights. Ammonium sulfate remained the most favorable salt precipitant, with a median concentration of 1.6 M. The crystallization pH for the vast majority of protein complexes was between 5.0 and 8.0. Overall, the boundaries for the crystallization configuration space of protein complexes appear to be more restricted than those of soluble proteins. This may reflect the limited stability and solubility of protein-protein complexes. Based on statistical analysis of the database, a sparse-matrix and a systematic buffer and pH screen were formulated to best represent the crystallization of protein complexes.

Buffers↗
PubMed2004

The human type I keratin gene family: characterization of new hair follicle specific members and evaluation of the chromosome 17q21.2 gene domain.

In general concurrence with recent studies, bioinformatic analysis of the chromosome 17q21.2 DNA sequence found in the EBI/Genebank database shows the presence of 27 type I keratin genes and five keratin pseudogenes present on 8 contiguous Bacterial Artificial Chromosome (BAC) sequences. This constitutes the 970 kb type I keratin gene domain. Inserted into this domain is a 350 kb region harboring 32 previously characterized keratin-associated protein genes. Of the 27 keratin genes found in this region, six have not been characterized in detail. This study reports the isolation of cDNA sequences for these keratin genes, termed K25irs1-K25irs4, Ka35, and Ka36, as well as cDNA sequences for the previously reported hair keratins hHa3-I, hHa7, and hHa8. RT-PCR analysis of 14 epithelial tissues using primers for the six novel keratins, as well as for keratins 23 and 24, shows that the six novel keratins appear to be hair follicle associated. Previous expression data, coupled with evolutionary analysis studies point to K25irs1-K25irs4 probably being inner root sheath specific keratins. Ka35 and Ka36 are, based on their exon-intron structure and expression characteristics, hair keratins. In contrast, K23 and K24 appear to be epithelial keratins associated with simple/glandular or stratified, non-cornified epithelia, respectively. A literature analysis coupled with the data presented here confirms that all of the 27 keratin genes found on this domain have been characterized at the transcriptional level. Together with K18, a type I keratin gene found on the type II keratin domain, this seems to be the entire complement of functional type I keratins in humans.

Amino Acid Sequence↗
PubMed2003

Mining single-nucleotide polymorphisms from hexaploid wheat ESTs.

Single-nucleotide polymorphisms (SNPs) represent a new form of functional marker, particularly when they are derived from expressed sequence tags (ESTs). A bioinformatics strategy was developed to discover SNPs within a large wheat EST database and to demonstrate the utility of SNPs in genetic mapping and genetic diversity applications. A collection of > 90000 wheat ESTs was assembled into contiguous sequences (contigs), and 45 random contigs were then visually inspected to identify primer pairs capable of amplifying specific alleles. We estimate that homoeologue sequence variants occurred 1 in 24 bp and the frequency of SNPs between wheat genotypes was 1 SNP/540 bp (theta = 0.0069). Furthermore, we estimate that one diagnostic SNP test can be developed from every contig with 10-60 EST members. Thus, EST databases are an abundant source of SNP markers. Polymorphism information content for SNPs ranged from 0.04 to 0.50 and ESTs could be mapped into a framework of microsatellite markers using segregating populations. The results showed that SNPs in wheat can be discovered in ESTs, validated, and be applied to conventional genetic studies.

Base Sequence↗
PubMed2006

Gene expression profiling of benign and malignant pheochromocytoma.

There are currently no reliable diagnostic and prognostic markers or effective treatments for malignant pheochromocytoma. This study used oligonucleotide microarrays to examine gene expression profiles in pheochromocytomas from 90 patients, including 20 with malignant tumors, the latter including metastases and primary tumors from which metastases developed. Other subgroups of tumors included those defined by tissue norepinephrine compared to epinephrine contents (i.e., noradrenergic versus adrenergic phenotypes), adrenal versus extra-adrenal locations, and presence of germline mutations of genes predisposing to the tumor. Correcting for the confounding influence of noradrenergic versus adrenergic catecholamine phenotype by the analysis of variance revealed a larger and more accurate number of genes that discriminated benign from malignant pheochromocytomas than when the confounding influence of catecholamine phenotype was not considered. Seventy percent of these genes were underexpressed in malignant compared to benign tumors. Similarly, 89% of genes were underexpressed in malignant primary tumors compared to benign tumors, suggesting that malignant potential is largely characterized by a less-differentiated pattern of gene expression. The present database of differentially expressed genes provides a unique resource for mapping the pathways leading to malignancy and for establishing new targets for treatment and diagnostic and prognostic markers of malignant disease. The database may also be useful for examining mechanisms of tumorigenesis and genotype-phenotype relationships. Further progress on the basis of this database can be made from follow-up confirmatory studies, application of bioinformatics approaches for data mining and pathway analyses, testing in pheochromocytoma cell culture and animal model systems, and retrospective and prospective studies of diagnostic markers.

Adrenal Gland Neoplasms↗
PubMed2003

Beyond the literature: bioinformatics training for medical librarians.

As genetics and molecular biology progressively impact on society and the practice of medicine, medical librarians must prepare themselves to deal with this new arena of information. The Eskind Biomedical Library at Vanderbilt University Medical Center has developed a bioinformatics training program for its librarians involving subject knowledge, literature evaluation, and database searching techniques. This program can serve as a model for other libraries.

Computational Biology↗
PubMed2002

Pharmacogenomics, regulation and signaling pathways of phase I and II drug metabolizing enzymes.

Drug or xenobiotics metabolizing enzymes (DMEs or XMEs) play central roles in the biotransformation, metabolism and/or detoxification of xenobiotics or foreign compounds, that are introduced to the human body. In general, DMEs protect or defend the body against the potential harmful insults from the environment. Once in the body, many xenobiotics may induce signal transduction events either specifically or non-specifically leading to various cellular, physiological and pharmacological responses including homeostasis, proliferation, differentiation, apoptosis, or necrosis. For the body to minimize the insults caused by these xenobiotics, various tissues/organs are well equipped with diverse DMEs including various Phase I and Phase II enzymes, which are present in abundance either at the basal level and/or increased/induced after exposure. To better understand the pharmacogenomic/gene expression profile of DMEs and the underlying molecular mechanisms after exposure to xenobiotics or drugs, we will review our current knowledge on DNA microarray technology in gene expression profiling and the signal transduction events elicited by various xenobiotics mediated by either specific receptors or non-specific signal transduction pathways. Pharmacogenomics is the study of genes and the gene products (proteins) essential for pharmacological or toxicological responses to pharmaceutical agents. In order to assess the battery of genes that are induced or repressed by xenobiotics and pharmaceutical agents, cDNA microarray or oligonucleotide-based DNA chip technology can be a powerful tool to analyze, simultaneously, the gene expression profiles that are induced or repressed by xenobiotics. The regulation of gene expression of the various phase I DMEs such as the cytochrome P450 (CYP) as well as phase II DMEs generally depends on the interaction of the xenobiotics with the receptors. For instance, the expression of CYP1 genes can be induced via the aryl hydrocarbon receptor (AhR) which dimerizes with the AhR nuclear translocator (ARNT), in response to many polycyclic aromatic hydrocarbon (PAHs). Similarly, the steroid family of orphan receptors, the constitutive androstane receptor (CAR) and pregnane X receptors (PXR), heterodimerize with the retinoid X receptor (RXR), transcriptionally activate the promoters of CYP2B and CYP3A gene expression by xenobiotics such as phenobarbital-like compounds (CAR) and dexamethasone and rifampin-type of agents (PXR). The peroxisome proliferator activated receptor (PPAR) which is one of the first characterized members of the nuclear hormone receptor, also dimerizes with RXR and it has been shown to be activated by lipid lowering agent fibrate-type of compounds leading to transcriptional activation of the promoters on the CYP4A genes. The transcriptional activation of these promoters generally leads to the induction of their mRNA. The physiological and the pharmacological implications of common partner of RXR for CAR, PXR, and PPAR receptors largely remain unknown and are under intense investigations. For the phase II DMEs, phase II gene inducers such as phenolic compounds butylated hydroxyanisol (BHA), tert-butylhydroquinone (tBHQ), green tea polyphenol (GTP), (-)-epicatechin-3-gallate (EGCG) and the isothiocyanates (PEITC, sulforaphane) generally appear to be electrophiles. They can activate the mitogen-activated protein kinase (MAPK) pathway via electrophilic-mediated stress response, resulting in the activation of bZIP transcription factors Nrf2 which dimerizes with Mafs and binds to the antioxidant/electrophile response element (ARE/EpRE) enhancers which are found in many phase II DMEs as well as many cellular defensive enzymes such as thioredoxins, gammaGCS and HO-1, with the subsequent induction of gene expression of these genes. It appears that in general, exposure to phase I or phase II gene inducers or xenobiotics may trigger a cellular "stress" response leading to the increase in the gene expression of these DMEs, which ultimately enhance the elimination and clearance of the xenobiotics e xenobiotics and/or the "cellular stresses" including harmful reactive intermediates such as reactive oxygen species (ROS), so that the body will remove the "stress" expeditiously. Consequently, this homeostatic response of the body plays a central role in the protection of the organism against environmental insults such as xenobiotics. Advances in DNA microarray technologies and mammalian genome sequencing will soon allow quantitative assessment of expression profiles of all genes in the selected tissues. The ability to predict phenotypic outcomes from gene expression profiles is currently in its infancy, however, and will require additional bioinformatic tools. Such tools will facilitate information gathering from literature and gene databases as well as integration of expression data with animal physiology studies. The study of pharmacogenomic/gene expression profile and the understanding of the regulation and the signal transduction mechanisms elicited by pharmaceutical agents can be of potential importance during drug discovery and the drug development.

Animals↗
PubMed2003

From immunome to vaccine: epitope mapping and vaccine design tools.

Since the publication of the complete genome of a pathogenic bacterium in 1995, more than 50 bacterial pathogens have been sequenced and at least 120 additional projects are currently underway. Faced with the expanding volume of information now available from genome databases, vaccinologists are turning to epitope mapping tools to screen vaccine candidates. Bioinformatics tools such as EpiMatrix and Conservatrix, which search for unique or multi-HLA-restricted (promiscuous) T cell epitopes and can find epitopes that are conserved across variant strains of the same pathogen, have accelerated the process of epitope mapping. Additional tools for screening epitopes for similarity to 'self' (BlastiMer) and for assembling putative epitopes into strings if they overlap (EpiAssembler) have been developed at EpiVax. Tools that map proteasome cleavage sires are available on the Internet. When used together, these bioinformatics tools offer a significant advantage over traditional methods of vaccine design since high throughput screening and design is performed in silico, followed by confirmatory studies in vitro. These new tools are being used to develop novel vaccines and therapeutics for the prevention and treatment of infectious diseases such as HIV, hepatitis C, tuberculosis, and some cancers. More recent applications of the tools involve deriving novel vaccine candidates directly from whole genomes, an approach that has been named 'genome to vaccine'.

Allergy and Immunology↗
PubMed2004

[Application of bioinformatics in researches of industrial biocatalysis].

Industrial biocatalysis is currently attracting much attention to rebuild or substitute traditional producing process of chemicals and drugs. One of key focuses in industrial biocatalysis is biocatalyst, which is usually one kind of microbial enzyme. In the recent, new technologies of bioinformatics have played and will continue to play more and more significant roles in researches of industrial biocatalysis in response to the waves of genomic revolution. One of the key applications of bioinformatics in biocatalysis is the discovery and identification of the new biocatalyst through advanced DNA and protein sequence search, comparison and analyses in Internet database using different algorithm and software. The unknown genes of microbial enzymes can also be simply harvested by primer design on the basis of bioinformatics analyses. The other key applications of bioinformatics in biocatalysis are the modification and improvement of existing industrial biocatalyst. In this aspect, bioinformatics is of great importance in both rational design and directed evolution of microbial enzymes. Based on the successful prediction of tertiary structures of enzymes using the tool of bioinformatics, the undermentioned experiments, i.e. site-directed mutagenesis, fusion protein construction, DNA family shuffling and saturation mutagenesis, etc, are usually of very high efficiency. On all accounts, bioinformatics will be an essential tool for either biologist or biological engineer in the future researches of industrial biocatalysis, due to its significant function in guiding and quickening the step of discovery and/or improvement of novel biocatalysts.

Biocatalysis↗
PubMed2006

Secured distributed service to manage biological data on EGEE grid.

Biological data are most times published and then become public ones. They, then, do not need to be isolated or encrypted. But, in some cases, these data stemed from patients or are analyzed with, for instance, pharmaceutical or agronomics goals. Also in simple ways , these data, before to become public, have to be kept confidential while researchers haven't been able to publish their work or to register them. So they are a lot of cases where the integrity and the confidentiality of biological data have to be protected against unauthorized accesses. But, as these private data are also large datasets, they need high-throughput computing and huge data storage to processed, such as ones produced by complete genome projects. These requirements are enhanced in the context of a Grid such EGEE, where the computing and storage resources are distributed across a large-scale platform. We have developed a secured distributed service to manage biological data on grid: the EncFile encrypted files management system. We have deployed it on the production platform of the EGEE grid project. Thus we provided grid users with a user-friendly component that doesn't require any user privileges. And we have integrated into a bioinformatics grid portal associated to encrypted representative biological resources: world-famous databases and programs.

Computational Biology↗
PubMed2003

Databases and data mining for computational vaccinology.

Drugs and vaccines are keys to the effective fight against disease. While the pharmaceutical industry has developed an awesome array of real and virtual approaches to rational drug discovery, the complexity of the immune system hampers attempts to design and develop vaccines in a rational manner. The goal of immunoinformatics (the application of informatics techniques to immunological macromolecules), an emergent sub-discipline of bioinformatics, is to develop computational vaccinology as a potent tool in the quest for new vaccines. Databases and data mining, the two principal weapons at the disposal of the in silico vaccinologist, will be presented in the light of current developments.

Computational Biology↗
PubMed2005

Searching for hypothetical proteins: theory and practice based upon original data and literature.

A large part of mammalian proteomes is represented by hypothetical proteins (HP), i.e. proteins predicted from nucleic acid sequences only and protein sequences with unknown function. Databases are far from being complete and errors are expected. The legion of HP is awaiting experiments to show their existence at the protein level and subsequent bioinformatic handling in order to assign proteins a tentative function is mandatory. Two-dimensional gel-electrophoresis with subsequent mass spectrometrical identification of protein spots is an appropriate tool to search for HP in the high-throughput mode. Spots are identified by MS or by MS/MS measurements (MALDI-TOF, MALDI-TOF-TOF) and subsequent software as e.g. Mascot or ProFound. In many cases proteins can thus be unambiguously identified and characterised; if this is not the case, de novo sequencing or Q-TOF analysis is warranted. If the protein is not identified, the sequence is being sent to databases for BLAST searches to determine identities/similarities or homologies to known proteins. If no significant identity to known structures is observed, the protein sequence is examined for the presence of functional domains (databases PROSITE, PRINTS, InterPro, ProDom, Pfam and SMART), subjected to searches for motifs (ELM) and finally protein-protein interaction databases (InterWeaver, STRING) are consulted or predictions from conformations are performed. We here provide information about hypothetical proteins in terms of protein chemical analysis, independent of antibody availability and specificity and bioinformatic handling to contribute to the extension/completion of protein databases and include original work on HP in the brain to illustrate the processes of HP identification and functional assignment.

Amino Acid Sequence↗