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At least 865 records · Page 48Linked to original sources
PubMed2006

A nifH-based oligonucleotide microarray for functional diagnostics of nitrogen-fixing microorganisms.

Nitrogen fixation is an important process in biogeochemical cycles exclusively carried out by prokaryotes, mostly by an evolutionarily conserved nitrogenase protein complex, of which one of the structural genes (nifH) is highly valuable for phylogenetic and diversity analyses. We developed a nifH-based short oligonucleotide microarray (nifH diagnostic microarray) as a rapid tool to effectively monitor nitrogen-fixing diazotrophic populations in a wide range of environments. Taking account of the overwhelming predominance of environmental nifH fragments from uncultivated microorganisms in public databases, our nifH microarray is mainly based on nifH sequences from as yet unidentified prokaryotes. Standard conditions for microarray performance were determined, and criteria for the design of specific oligonucleotides were defined. A primary set of 56 oligonucleotides was validated with fluorescence-labeled single-stranded nifH targets from five reference strains, 26 environmental clones, and artificial mixtures of reference strains. The nifH microarray was applied to analyze the diversity (based on DNA) and activity (based on mRNA) of diazotrophs in roots of wild rice samples from Namibia. Results demonstrated that only a small subset of diazotrophs being present in the sample were actually fixing nitrogen actively. Our data suggest that the developed nifH microarray is a highly reproducible and semiquantitative method for mapping the variability of diazotrophic diversity, allowing rapid comparisons of the relative abundance and activity of diazotrophic prokaryotes in the environment. A further refined nifH microarray comprising of 194 oligonucleotide probes now covers more than 90% of sequences in our nifH database.

Azoarcus↗
PubMed2002

A novel type of conserved DNA-binding domain in the transcriptional regulators of the AlgR/AgrA/LytR family.

Sequence analysis of bacterial genomes revealed a novel DNA-binding domain. This domain is found in several response regulators of the two-component signal transduction system, such as Pseudomonas aeruginosa AlgR, involved in the regulation of alginate biosynthesis and in the pathogenesis of cystic fibrosis; Clostridium perfringens VirR, a regulator of virulence factors, and in several regulators of bacteriocin biosynthesis, previously unified in the AgrA/ComE family. Most of the transcriptional regulators that contain this DNA-binding domain are involved in biosynthesis of extracellular polysaccharides, fimbriation, expression of exoproteins, including toxins, and quorum sensing. We refer to it as the LytTR ('litter') domain, after Bacillus subtilis LytT and Staphylococcus aureus LytR response regulators, involved in regulation of cell autolysis. In addition to response regulators, the LytTR domain is found in combination with MHYT, PAS and other sensor domains.

Amino Acid Sequence↗
PubMedDate not supplied

Reproducibility of polypeptide spot positions in two-dimensional gels run using carrier ampholytes in the isoelectric focusing dimension.

The reproducibility of complex protein patterns in two-dimensional (2-D) gels run with carrier ampholytes in the first dimension has been investigated. Two different laboratories collaborated in the study and 18 or 19 gels were run in each laboratory for comparison. The electrophoresis chemicals, running devices, and samples were standardized in both labs. The resulting 37 gels were scanned with a charge-coupled device (CCD) camera and spots were located, counted, quantified, and matched using a commercially available image analysis system. Subsequently, the reproducibility of spot position was determined. To perform the statistical analysis, the test gels were initially each matched to a master reference gel. Next, three sets of 12 gels (the image analysis software database could analyze only 12 gels at a time) were analyzed and the isoelectric point (pI) and molecular weight (M(r)) positional variation of all the spots that matched across the gels in each set was determined. The resulting statistical analysis indicates very high reproducibility of the carrier ampholyte technique.

Buffers↗
PubMed2007

Critical evaluation of methods to incorporate entropy loss upon binding in high-throughput docking.

Proper accounting of the positional/orientational/conformational entropy loss associated with protein-ligand binding is important to obtain reliable predictions of binding affinity. Herein, we critically examine two simplified statistical mechanics-based approaches, namely a constant penalty per rotor method, and a more rigorous method, referred to here as the partition function-based scoring (PFS) method, to account for such entropy losses in high-throughput docking calculations. Our results on the estrogen receptor beta and dihydrofolate reductase proteins demonstrate that, while the constant penalty method over-penalizes molecules for their conformational flexibility, the PFS method behaves in a more "DeltaG-like" manner by penalizing different rotors differently depending on their residual entropy in the bound state. Furthermore, in contrast to no entropic penalty or the constant penalty approximation, the PFS method does not exhibit any bias towards either rigid or flexible molecules in the hit list. Preliminary enrichment studies using a lead-like random molecular database suggest that an accurate representation of the "true" energy landscape of the protein-ligand complex is critical for reliable predictions of relative binding affinities by the PFS method.

Algorithms↗
PubMed1994

MHCDB--database of the human MHC.

Genetic and physical data relating to the human major histocompatibility complex (MHC) were compiled and analyzed, using a genome analysis program. The current contents of the database include: 1) location of over 100 genes and other markers; 2) location of over 250 YAC and cosmid clones; 3) 150 kilobases of genomic DNA sequence including full annotation (exon/intron boundaries, repeats, promoters, etc.); 4) cDNA sequences of currently-known class I and class II alleles; and 5) accompanying descriptive data--references, comments, laboratory addresses, and so on. The database allows rapid access, retrieval, and display of all these data, which should make it a useful tool for the study of the human MHC. MHCDB is publicly available and will be updated as new data become available.

Alleles↗
PubMed2004

Membrane-associated and secreted genes in breast cancer.

The identification of membrane-associated and secreted genes that are differentially expressed is a useful step in defining new targets for the diagnosis and treatment of cancer. Extracting information on the subcellular localization of genes represented on DNA microarrays is difficult and is limited by the incomplete sequence and annotation that is available in existing databases. Here we combine a biochemical and bioinformatic approach to identify membrane-associated and secreted genes expressed in the MCF-7 breast cancer cell line. Our approach is based on the analysis of differential hybridization levels of RNAs that have been physically separated by virtue of their association with polysomes on the endoplasmic reticulum. This approach is specifically applicable to oligonucleotide microarrays such as Affymetrix, which use single-color hybridization instead of dual-color competitive hybridizations. Assignment to membrane-associated and secreted class membership is based on both the differential hybridization levels and an expression threshold, which are calculated empirically from data collected on a reference set of known cytoplasmic and membrane proteins. This method enabled the identification of 755 membrane-associated and secreted probe sets expressed in MCF-7 cells for which this annotation did not previously exist. The data were used to filter a previously reported expression dataset to identify membrane-associated and secreted genes which are associated with poor prognosis in breast cancer and represent potential targets for diagnosis and treatment. The approach reported here should provide a useful tool for the analysis of gene expression patterns, identifying membrane-associated or secreted genes with biological relevance that have the potential for clinical applications in diagnosis or treatment.

Breast Neoplasms↗
PubMed1999

Proteome map of the human hippocampus.

The proteins expressed by a genome have been termed the proteome. By comparing the proteome of a disease-affected tissue with the proteome of an unaffected tissue it is possible to identify proteins that play a role in a disease process. The hippocampus is involved in the processing of short-term memory and is affected in Alzheimer's disease. Any comparative proteome analysis that can identify proteins important in a disease affecting the hippocampus requires the characterization of the normal hippocampal proteome. Therefore, we homogenised normal hippocampal tissue and separated the proteins by two-dimensional polyacrylamide gel electrophoresis (2DE). Seventy-two unique protein spots were collected from Coomassie blue-stained 2DE gels and subjected to in-gel digestion with trypsin, reversed-phase high-pressure liquid chromatography peptide separation, and N-terminal protein sequencing. Sufficient protein sequence was obtained to successfully characterize 66 of the 72 protein spots chosen (92%). Three of the 66 proteins were not present in any database (4.5%). The characterized proteins comprised two dominant functional groups, i.e., enzymes involved in intermediary cellular metabolism (40%), and proteins associated with the cytoskeleton (15%). The identity, molecular mass, isoelectric point, and relative concentration of the characterized proteins are described and constitute a partial proteome map of the normal human hippocampus.

Adult↗
PubMed2001

Evaluating the potential of using fold-recognition models for molecular replacement.

Here, the proposal is investigated that protein tertiary structure prediction methods and threading methods in particular might be applied to the problem of solving a protein structure by X-ray crystallography, thus reducing the need for the more traditional experimental intensity methods of data phasing, such as heavy-metal isomorphous replacement and anomalous scattering methods, and without reference to a very closely related protein of known structure. If this kind of approach were to become successful and reliable, this would represent a significant advance in protein structure determination, offering an easy and accessible method for the initial data phasing for proteins' crystal structures, utilizing the vast amount of structural data, deposited in the Brookhaven PDB, that has been accumulated over the past 30 years of crystallographic structural studies. In the light of the ongoing structural genomics initiatives, the successful development of this kind of approach would be of enormous benefit.

Crystallography, X-Ray↗
PubMed2003

Mapping the proteome of barrel medic (Medicago truncatula).

A survey of six organ-/tissue-specific proteomes of the model legume barrel medic (Medicago truncatula) was performed. Two-dimensional polyacrylamide gel electrophoresis reference maps of protein extracts from leaves, stems, roots, flowers, seed pods, and cell suspension cultures were obtained. Five hundred fifty-one proteins were excised and 304 proteins identified using peptide mass fingerprinting and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Nanoscale high-performance liquid chromatography coupled with tandem quadrupole time-of-flight mass spectrometry was used to validate marginal matrix-assisted laser desorption ionization time-of-flight mass spectrometry protein identifications. This dataset represents one of the most comprehensive plant proteome projects to date and provides a basis for future proteome comparison of genetic mutants, biotically and abiotically challenged plants, and/or environmentally challenged plants. Technical details concerning peptide mass fingerprinting, database queries, and protein identification success rates in the absence of a sequenced genome are reported and discussed. A summary of the identified proteins and their putative functions are presented. The tissue-specific expression of proteins and the levels of identified proteins are compared with their related transcript abundance as quantified through EST counting. It is estimated that approximately 50% of the proteins appear to be correlated with their corresponding mRNA levels.

Amino Acid Sequence↗
PubMed2006

Multiple predictors of coronary restenosis after drug-eluting stent implantation in patients with diabetes.

OBJECTIVES: To identify parameters influencing the likelihood of restenosis after implantation of drug-eluting stents (DES) in patients with diabetes. METHODS: Stented patients (n = 840) with DES were retrospectively reviewed for inclusion in the study from the Multicenter PCI Database Registry. From this database, 211 (25.1%) of 840 patients with six-month angiographic follow up had diabetes. Predictors of coronary restenosis were identified with univariate and multivariate logistic regression analyses. RESULTS: Restenosis occurred in 92 of 629 (14.6%) patients without diabetes and in 44 (20.9%) of 211 patients with diabetes (p < 0.001). Multivariate parameters for predicting restenosis in the diabetic group were current smoking (odds ratio (OR) 1.923, 95% confidence interval (CI) 1.055 to 4.725, p = 0.036), higher C reactive protein concentration (OR 1.031, 95% CI 1.011 to 1.075, p = 0.043), use of the paclitaxel-eluting stent (OR 2.638, 95% CI 1.338 to 5.200, p = 0.005), longer stent length (OR 1.065, 95% CI 1.021 to 1.119, p = 0.033), smaller reference diameter before DES implantation (OR 0.501, 95% CI 0.110 to 0.965, p = 0.040), smaller reference diameter (OR 0.455, 95% CI 0.120 to 0.814, p = 0.026) and minimum lumen diameter (OR 0.447, 95% CI 0.068 to 0.876, p = 0.039) after DES implantation. CONCLUSION: Even with the introduction of DES, diabetes remains a significant predictor of coronary restenosis, especially in cases of a small baseline vessel size, small vessel size after percutaneous coronary intervention, longer stent length, use of the paclitaxel-eluting stent, current smoking and high C reactive protein concentration.

Angioplasty, Balloon↗
PubMed1998

Correlation of acidic and basic carrier ampholyte and immobilized pH gradient two-dimensional gel electrophoresis patterns based on mass spectrometric protein identification.

Separation of proteins on either carrier ampholyte-based or immobilized pH gradient-based two-dimensional (2-D) gels gives rise to electrophoretic patterns that are difficult to compare visually. In this paper we have used matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to determine the identities of 335 protein spots in these two 2-D gel systems, including a substantial number of basic proteins which had never been identified before. Proteins that were identified in both gel systems allowed us to cross-reference the gel patterns. Vector analysis of these cross-references demonstrated that there is no obvious pattern by which the mobility of a protein in one gel system can be used to predict its mobility in the other. Thus, as laboratories adopt the immobilized pH gradient-based 2-D gel systems, the only reliable means of translating the data gained with the carrier ampholyte-based gel system is to positively identify the proteins in both 2-D systems.

Ampholyte Mixtures↗
PubMed1989

ALADDIN: an integrated tool for computer-assisted molecular design and pharmacophore recognition from geometric, steric, and substructure searching of three-dimensional molecular structures.

ALADDIN is a computer program for the design or recognition of compounds that meet geometric, steric, and substructural criteria. ALADDIN searches a database of three-dimensional structures, marks atoms that meet substructural criteria, evaluates geometric criteria, and prepares a number of files that are input for molecular modification and coordinate generation as well as for molecular graphics. Properties calculated from the three-dimensional structure are described by either properties calculated from the molecule itself or from the molecule as compared to a reference molecule and associated surfaces. ALADDIN was used to design analogues to probe a bioactive conformation of a small molecule and a peptide, to test alternative superposition rules for receptor mapping of the D2 dopamine receptor, to recognize unexpected D2 dopamine agonist activity of existing compounds, and to design compounds to fit a binding site on a protein of known structure. We have found that series designed by ALADDIN show much more subtle variation in shape than do those designed by traditional methods and that compounds can be designed to be very close matches to the objective.

Binding Sites↗
PubMed2002

Familial abdominal aortic aneurysm: a systematic review of a genetic background.

BACKGROUND: Familial clustering of the abdominal aortic aneurysm (AAA) is clear, 12-19% of AAA patients have one or more first-degree relatives with an aneurysm and 4-19% is detected with ultrasound screening. OBJECTIVES: To review the genetic background of AAA. DESIGN, METHODS AND MATERIALS: Computer searches of the MEDLINE, EMBASE, SUMsearch database and the Cochrane Library and searched reference lists of English language articles concerning the genetics of AAA, candidate gene approach and linkage analysis. RESULTS: Brothers of AAA patients are at high risk to develop an AAA. The candidate gene approach was performed to detect defects in one of the components of the connective tissue, i.e. type I and III collagen, elastin and fibrillin, the inflammatory cell-derived matrix metalloproteinase, there inhibitors, auto-immune components and components related to atherosclerosis. CONCLUSION: These studies give us insight in the pathology but do not lead to the specific genetic factor(s) responsible for (familial) AAA. Considering the supposed autosomal dominant inheritance, a gene mutation in one of the structural proteins of the connective tissue is expected. In the future, linkage analysis may resolve the genetic background of AAA.

Aortic Aneurysm, Abdominal↗
PubMed1997

Development of a cell signaling networks database.

In multicellular organisms cell signaling networks play important roles in wide range of biological phenomena, such as development, differentiation, reproduction, morphogenesis, carcinogenesis, apoptosis, and even learning. In order to explain biological phenomena based on cell signaling models, we have developed a database for cell signaling networks. The system contains mechanisms of signal transduction and structure and functional data and references of extracellular chemicals and biomolecules. CSNDB is constructed using ACEDB system, and includes various graphical representations such as pathway diagrams, map diagrams, 3-D images, pictures, and VRML environment. The system will be useful for modeling cells and their information processing, and to explain important biological phenomena based on these models.

Cell Cycle↗
PubMedDate not supplied

Computer identification of Shigella species by rRNA gene restriction patterns.

We describe a MluI ribotyping scheme for Shigella which approaches correlation with serotyping. One hundred and seventeen reference strains and previously serotyped clinical isolates representing the 57 Shigella serotypes and biotypes were included in this study. A total of 51 distinct ribotypes were obtained and a database was built with them. The number of bands composing each ribotype varied from 9 to 15. The fragments ranged in size from 1.6 to 18.8 kbp. One hundred and eleven clinical isolates were successfully identified in a double blind study with standard biochemical/serologic methods, by automatic comparison of their ribotypes with our database using the software Taxotron.

Bacterial Proteins↗
PubMed2005

Molecular models of NS3 protease variants of the Hepatitis C virus.

BACKGROUND: Hepatitis C virus (HCV) currently infects approximately three percent of the world population. In view of the lack of vaccines against HCV, there is an urgent need for an efficient treatment of the disease by an effective antiviral drug. Rational drug design has not been the primary way for discovering major therapeutics. Nevertheless, there are reports of success in the development of inhibitor using a structure-based approach. One of the possible targets for drug development against HCV is the NS3 protease variants. Based on the three-dimensional structure of these variants we expect to identify new NS3 protease inhibitors. In order to speed up the modeling process all NS3 protease variant models were generated in a Beowulf cluster. The potential of the structural bioinformatics for development of new antiviral drugs is discussed. RESULTS: The atomic coordinates of crystallographic structure 1CU1 and 1DY9 were used as starting model for modeling of the NS3 protease variant structures. The NS3 protease variant structures are composed of six subdomains, which occur in sequence along the polypeptide chain. The protease domain exhibits the dual beta-barrel fold that is common among members of the chymotrypsin serine protease family. The helicase domain contains two structurally related beta-alpha-beta subdomains and a third subdomain of seven helices and three short beta strands. The latter domain is usually referred to as the helicase alpha-helical subdomain. The rmsd value of bond lengths and bond angles, the average G-factor and Verify 3D values are presented for NS3 protease variant structures. CONCLUSIONS: This project increases the certainty that homology modeling is an useful tool in structural biology and that it can be very valuable in annotating genome sequence information and contributing to structural and functional genomics from virus. The structural models will be used to guide future efforts in the structure-based drug design of a new generation of NS3 protease variants inhibitors. All models in the database are publicly accessible via our interactive website, providing us with large amount of structural models for use in protein-ligand docking analysis.

Amino Acid Sequence↗
PubMed2005

Sesame allergy: a growing food allergy of global proportions?

OBJECTIVE: To present an up-to-date synopsis on the global prevalence, natural history, nature of allergens, and immune mechanisms of sesame allergy. DATA SOURCES: We performed a literature search using the PubMed database. We obtained information on regulatory issues from the Web pages of respective agencies. STUDY SELECTION: The PubMed search was performed using the algorithm sesame AND allergy. Date restriction was not used, and only articles in English were considered. The articles obtained were screened for additional references to work not obtained in the initial search. Each article was analyzed, and a pooled source of published information was generated. RESULTS: Evidence was found for increased reporting of sesame allergy during the past 5 decades, with reports mostly from developed countries. Clinically, most sesame allergy was presented in at least 2 major forms: (1) immediate hypersensitivity, often expressed as systemic anaphylaxis, associated with positive skin prick test and/or IgE antibody test results to sesame proteins with some cross-reactivity with other foods, and (2) delayed hypersensitivity to lignin-like compounds in sesame oil clinically expressed as contact allergic dermatitis. There were a few cases of immediate hypersensitivity to sesame with negative skin prick test and/or IgE antibody test results that were confirmed by oral challenge tests. CONCLUSIONS: Sesame allergy is a significant, serious, and growing problem. Evidence exists of the ability of protein and oil components of sesame to trigger immediate hypersensitivity via IgE antibody and delayed hypersensitivity via cell-mediated immune responses, respectively.

Allergens↗
PubMed1999

Universal protein families and the functional content of the last universal common ancestor.

The phylogenetic distribution of Methanococcus jannaschii proteins can provide, for the first time, an estimate of the genome content of the last common ancestor of the three domains of life. Relying on annotation and comparison with reference to the species distribution of sequence similarities results in 324 proteins forming the universal family set. This set is very well characterized and relatively small and nonredundant, containing 301 biochemical functions, of which 246 are unique. This universal function set contains mostly genes coding for energy metabolism or information processing. It appears that the Last Universal Common Ancestor was an organism with metabolic networks and genetic machinery similar to those of extant unicellular organisms.

Archaeal Proteins↗