Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Genomic database”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,675 records · Page 93Linked to original sources
PubMed2005

Identification and targeted disruption of the gene encoding the main 3-ketosteroid dehydrogenase in Mycobacterium smegmatis.

The catabolic potential for sterol degradation of fast-growing mycobacteria is well known. However, no genes or enzymes responsible for the steroid degradation process have been identified as yet in these species. One of the key enzymes required for degradation of the steroid ring structure is 3-ketosteroid Delta(1)-dehydrogenase (KsdD). The recent annotation of the Mycobacterium smegmatis genome (TIGR database) revealed six KsdD homologues. Targeted disruption of the MSMEG5898 (ksdD-1) gene, but not the MSMEG4855 (ksdD-2) gene, resulted in partial inactivation of the cholesterol degradation pathway and accumulation of the intermediate 4-androstene-3,17-dione. This effect was reversible by the introduction of the wild-type ksdD-1 gene into M. smegmatis DeltaksdD-1 or overexpression of ksdD-2. The data indicate that KsdD1 is the main KsdD in M. smegmatis, but that KsdD2 is able to perform the cholesterol degradation process when overproduced.

Cloning, Molecular↗
PubMed2000

Molecular cloning, genomic organization, developmental regulation, and a knock-out mutant of a novel leu-rich repeats-containing G protein-coupled receptor (DLGR-2) from Drosophila melanogaster.

After screening the Berkeley Drosophila Genome Project database with sequences from a recently characterized Leu-rich repeats-containing G protein-coupled receptor (LGR) from Drosophila (DLGR-1), we identified a second gene for a different LGR (DLGR-2) and cloned its cDNA. DLGR-2 is 1360 amino acid residues long and shows a striking structural homology with members of the glycoprotein hormone [thyroid-stimulating hormone (TSH); follicle-stimulating hormone (FSH); luteinizing hormone/choriogonadotropin (LH/CG)] receptor family from mammals and with two additional, recently identified mammalian orphan LGRs (LGR-4 and LGR-5). This homology includes the seven transmembrane region (e.g., 49% amino acid identity with the human TSH receptor) and the very large extracellular amino terminus. This amino terminus contains 18 Leu-rich repeats-in contrast with the 3 mammalian glycoprotein hormone receptors and DLGR-1 that contain 9 Leu-rich repeats, but resembling the mammalian LGR-4 and LGR-5 that each have 17 Leu-rich repeats in their amino termini. The DLGR-2 gene is >18.6 kb pairs long and contains 15 exons and 14 introns. Four intron positions coincide with the intron positions of the three mammalian glycoprotein hormone receptors and have the same intron phasing, showing that DLGR-2 is evolutionarily related to these mammalian receptors. The DLGR-2 gene is located at position 34E-F on the left arm of the second chromosome and is expressed in embryos and pupae but not in larvae and adult flies. Homozygous knock-out mutants, where the DLGR-2 gene is interrupted by a P element insertion, die around the time of hatching. This finding, together with the expression data, strongly suggests that DLGR-2 is exclusively involved in development.

Alternative Splicing↗
PubMed2006

GeneDesign: rapid, automated design of multikilobase synthetic genes.

Modern molecular biology has brought many new tools to the geneticist as well as an exponentially expanding database of genomes and new genes for study. Of particular use in the analysis of these genes is the synthetic gene, a nucleotide sequence designed to the specifications of the investigator. Typically, synthetic genes encode the same product as the gene of interest, but the synthetic nucleotide sequence for that protein may contain modifications affecting expression or base composition. Other desirable changes typically involve the revision of restriction sites. Designing synthetic genes by hand is a time-consuming and error-prone process that may involve several computer programs. We have developed a tools environment that combines many modules to provide a platform for rapid synthetic gene design for multikilobase sequences. We have used GeneDesign to successfully design a synthetic Ty1 element and a large variety of other synthetic sequences. GeneDesign has been implemented as a publicly accessible Web-based resource and can be found at http://slam.bs.jhmi.edu/gd.

Genetic Engineering↗
PubMed2003

Identification of AtNDI1, an internal non-phosphorylating NAD(P)H dehydrogenase in Arabidopsis mitochondria.

Plant mitochondria contain non-phosphorylating NAD(P)H dehydrogenases (DHs) that are not found in animal mitochondria. The physiological function, substrate specificity, and location of enzymes within this family have yet to be conclusively determined. We have linked genome sequence information to protein and biochemical data to identify that At1g07180 (SwissProt Q8GWA1) from the Arabidopsis Genome Initiative database encodes AtNDI1, an internal NAD(P)H DH in Arabidopsis mitochondria. Three lines of evidence are presented: (a). The predicted protein sequence of AtNDI1 has high homology with other designated NAD(P)H DHs from microorganisms, (b). the capacity for matrix NAD(P)H oxidation via the rotenone-insensitive pathway is significantly reduced in the Atndi1 mutant plant line, and (c). the in vitro translation product of AtNDI1 is imported into isolated mitochondria and located on the inside of the inner membrane.

Amino Acid Sequence↗
PubMed2005

Conservation of Arabidopsis flowering genes in model legumes.

The model plants Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa) have provided a wealth of information about genes and genetic pathways controlling the flowering process, but little is known about the corresponding pathways in legumes. The garden pea (Pisum sativum) has been used for several decades as a model system for physiological genetics of flowering, but the lack of molecular information about pea flowering genes has prevented direct comparison with other systems. To address this problem, we have searched expressed sequence tag and genome sequence databases to identify flowering-gene-related sequences from Medicago truncatula, soybean (Glycine max), and Lotus japonicus, and isolated corresponding sequences from pea by degenerate-primer polymerase chain reaction and library screening. We found that the majority of Arabidopsis flowering genes are represented in pea and in legume sequence databases, although several gene families, including the MADS-box, CONSTANS, and FLOWERING LOCUS T/TERMINAL FLOWER1 families, appear to have undergone differential expansion, and several important Arabidopsis genes, including FRIGIDA and members of the FLOWERING LOCUS C clade, are conspicuously absent. In several cases, pea and Medicago orthologs are shown to map to conserved map positions, emphasizing the closely syntenic relationship between these two species. These results demonstrate the potential benefit of parallel model systems for an understanding of flowering phenology in crop and model legume species.

Arabidopsis↗
PubMed2004

Functional properties of the protein disulfide oxidoreductase from the archaeon Pyrococcus furiosus: a member of a novel protein family related to protein disulfide-isomerase.

Protein disulfide oxidoreductases are ubiquitous redox enzymes that catalyse dithiol-disulfide exchange reactions with a CXXC sequence motif at their active site. A disulfide oxidoreductase, a highly thermostable protein, was isolated from Pyrococcus furiosus (PfPDO), which is characterized by two redox sites (CXXC) and an unusual molecular mass. Its 3D structure at high resolution suggests that it may be related to the multidomain protein disulfide-isomerase (PDI), which is currently known only in eukaryotes. This work focuses on the functional characterization of PfPDO as well as its relation to the eukaryotic PDIs. Assays of oxidative, reductive, and isomerase activities of PfPDO were performed, which revealed that the archaeal protein not only has oxidative and reductive activity, but also isomerase activity. On the basis of structural data, two single mutants (C35S and C146S) and a double mutant (C35S/C146S) of PfPDO were constructed and analyzed to elucidate the specific roles of the two redox sites. The results indicate that the CPYC site in the C-terminal half of the protein is fundamental to reductive/oxidative activity, whereas isomerase activity requires both active sites. In comparison with PDI, the ATPase activity was tested for PfPDO, which was found to be cation-dependent with a basic pH optimum and an optimum temperature of 90 degrees C. These results and an investigation on genomic sequence databases indicate that PfPDO may be an ancestor of the eukaryotic PDI and belongs to a novel protein disulfide oxidoreductase family.

Adenosine Triphosphate↗
PubMed1992

Gene structure of Bombyx mori larval serum protein (BmLSP).

To understand the molecular mechanisms of the larval-specific transcription of Bombyx mori larval serum protein (BmLSP), we isolated a clone of the BmLSP gene from a genomic library and sequenced a 3.5-kb fragment. An intron was found in the 5' noncoding region of the BmLSP gene. A putative transcription start point was determined by primer extension analysis. Genomic Southern hybridization showed that there is one copy of the BmLSP gene in a haploid genome. A database search revealed that the BmLSP gene has presumptive repetitive sequences found in other B. mori genes, the sequence homologous to ecdysone-responsive elements and a heptamer sequence found in storage protein genes.

Amino Acid Sequence↗
PubMed2004

The role of the ESSS protein in the assembly of a functional and stable mammalian mitochondrial complex I (NADH-ubiquinone oxidoreductase).

The ESSS protein is a recently identified subunit of mammalian mitochondrial complex I. It is a relatively small integral membrane protein (122 amino acids) found in the beta-subcomplex. Genomic sequence database searches reveal its localization to the X-chromosome in humans and mouse. The ESSS cDNA from Chinese hamster cells was cloned and shown to complement one complementation group of our previously described mutants with a proposed X-linkage. Sequence analyses of the ESSS cDNA in these mutants revealed chain termination mutations. In two of these mutants the protein is truncated at the C-terminus of the targeting sequence; the mutants are null mutants for the ESSS subunit. There is no detectable complex I assembly and activity in the absence of the ESSS subunit as revealed by blue native polyacrylamide gel electrophoresis (BN/PAGE) analysis and polarography. Complex I activity can be restored with ESSS subunits tagged with either hemagglutinin (HA) or hexahistidine (His6) epitopes at the C-terminus. Although, the accumulation of ESSS-HA is not dependent upon the presence of mtDNA-encoded subunits (ND1-6,4 L), it is incorporated into complex I only in presence of compatible complex I subunits from the same species.

Amino Acid Sequence↗
PubMed2000

Aldolases of the DhnA family: a possible solution to the problem of pentose and hexose biosynthesis in archaea.

Sequence analysis of the recently identified class I aldolase of Escherichia coli (dhnA gene product) helped to identify its homologs in Chlamydia trachomatis, Chlamydiophyla pneumoniae and in each of the completely sequenced archaeal genomes. Iterative database searches revealed sequence similarities between the DhnA-family enzymes, deoxyribose phosphate aldolases and bacterial (class II) fructose bisphosphate aldolases and allowed prediction of similar three-dimensional structures (TIM-barrel fold) in all these enzymes. The Schiff base-forming lysyl residues of DhnA and deoxyribose phosphate aldolase are conserved in all members of the DhnA and deoxyribose phosphate aldolase families, indicating that these enzymes share common features with both class I and class II aldolases. The DhnA-family enzymes are predicted to possess an aldolase activity and to play a critical role in sugar biosynthesis in archaea.

Amino Acid Sequence↗
PubMed2005

Bioinformatic and enzymatic characterization of the MAPEG superfamily.

The membrane associated proteins in eicosanoid and glutathione metabolism (MAPEG) superfamily includes structurally related membrane proteins with diverse functions of widespread origin. A total of 136 proteins belonging to the MAPEG superfamily were found in database and genome screenings. The members were found in prokaryotes and eukaryotes, but not in any archaeal organism. Multiple sequence alignments and calculations of evolutionary trees revealed a clear subdivision of the eukaryotic MAPEG members, corresponding to the six families of microsomal glutathione transferases (MGST) 1, 2 and 3, leukotriene C4 synthase (LTC4), 5-lipoxygenase activating protein (FLAP), and prostaglandin E synthase. Prokaryotes contain at least two distinct potential ancestral subfamilies, of which one is unique, whereas the other most closely resembles enzymes that belong to the MGST2/FLAP/LTC4 synthase families. The insect members are most similar to MGST1/prostaglandin E synthase. With the new data available, we observe that fish enzymes are present in all six families, showing an early origin for MAPEG family differentiation. Thus, the evolutionary origins and relationships of the MAPEG superfamily can be defined, including distinct sequence patterns characteristic for each of the subfamilies. We have further investigated and functionally characterized representative gene products from Escherichia coli, Synechocystis sp., Arabidopsis thaliana and Drosophila melanogaster, and the fish liver enzyme, purified from pike (Esox lucius). Protein overexpression and enzyme activity analysis demonstrated that all proteins catalyzed the conjugation of 1-chloro-2,4-dinitrobenzene with reduced glutathione. The E. coli protein displayed glutathione transferase activity of 0.11 micromol.min(-1).mg(-1) in the membrane fraction from bacteria overexpressing the protein. Partial purification of the Synechocystis sp. protein yielded an enzyme of the expected molecular mass and an N-terminal amino acid sequence that was at least 50% pure, with a specific activity towards 1-chloro-2,4-dinitrobenzene of 11 micromol.min(-1).mg(-1). Yeast microsomes expressing the Arabidopsis enzyme showed an activity of 0.02 micromol.min(-1).mg(-1), whereas the Drosophila enzyme expressed in E. coli was highly active at 3.6 micromol.min(-1).mg(-1). The purified pike enzyme is the most active MGST described so far with a specific activity of 285 micromol.min(-1).mg(-1). Drosophila and pike enzymes also displayed glutathione peroxidase activity towards cumene hydroperoxide (0.4 and 2.2 micromol.min(-1).mg(-1), respectively). Glutathione transferase activity can thus be regarded as a common denominator for a majority of MAPEG members throughout the kingdoms of life whereas glutathione peroxidase activity occurs in representatives from the MGST1, 2 and 3 and PGES subfamilies.

Animals↗
PubMed2002

RNA silencing: the genome's immune system.

Genomes are databases sensitive to invasion by viruses. In recent years, a defense mechanism has been discovered, which turns out to be conserved among eukaryotes. The system can be compared to the immune system in several ways: It has specificity against foreign elements and the ability to amplify and raise a massive response against an invading nucleic acid. The latter property is beginning to be understood at the molecular level.

Animals↗
PubMed2005

Protein structures forming the shell of primitive bacterial organelles.

Bacterial microcompartments are primitive organelles composed entirely of protein subunits. Genomic sequence databases reveal the widespread occurrence of microcompartments across diverse microbes. The prototypical bacterial microcompartment is the carboxysome, a protein shell for sequestering carbon fixation reactions. We report three-dimensional crystal structures of multiple carboxysome shell proteins, revealing a hexameric unit as the basic microcompartment building block and showing how these hexamers assemble to form flat facets of the polyhedral shell. The structures suggest how molecular transport across the shell may be controlled and how structural variations might govern the assembly and architecture of these subcellular compartments.

Amino Acid Sequence↗
PubMed2004

Saccharomyces cerevisiae JEN1 promoter activity is inversely related to concentration of repressing sugar.

When carbon sources are changed, Saccharomyces cerevisiae transcriptional patterns drastically change. To identify genes whose transcription can be used to quantitatively measure sugar concentrations, we searched genomic expression databases for a set of genes that are highly induced during the diauxic shift, and we used the promoters from these genes to drive expression of green fluorescent protein (GFP). Certain sugars, including glucose, fructose, and mannose, repress the promoter of JEN1, which encodes a lactate-pyruvate transporter, in a dose-dependent manner. Nonrepressing carbon sources include galactose, raffinose, ethanol, lactate, and glycerol. JEN1 promoter activity is a linear function of glucose concentration when organisms are grown at a steady-state glucose concentration below 1 g/liter. JEN1 promoter repression is specific to carbon source; heat or cold shock, osmotic stress, DNA damage, and nitrogen starvation do not significantly affect promoter activity. Activation of the JEN1 promoter requires the Snf1 protein kinase, but multiple regulatory elements most likely combine to provide the linear relationship between JEN1 promoter activity and sugar concentration. Thus, a JEN1 promoter-reporter system appears to provide a good living cell biosensor for the concentration of certain sugars. The JEN1 promoter also permits quantitative regulation of cellular functions not normally controlled by sugar concentrations. For example, a strain expressing FLO1 under control of the JEN1 promoter flocculates at a low glucose concentration.

Bacterial Proteins↗
PubMed2005

Regulation of resistance to copper in Xanthomonas axonopodis pv. vesicatoria.

Copper-resistant strains of Xanthomonas axonopodis pv. vesicatoria were previously shown to carry plasmid-borne copper resistance genes related to the cop and pco operons of Pseudomonas syringae and Escherichia coli, respectively. However, instead of the two-component (copRS and pcoRS) systems determining copper-inducible expression of the operons in P. syringae and E. coli, a novel open reading frame, copL, was found to be required for copper-inducible expression of the downstream multicopper oxidase copA in X. axonopodis. copL encodes a predicted protein product of 122 amino acids that is rich in histidine and cysteine residues, suggesting a possible direct interaction with copper. Deletions or frameshift mutations within copL, as well as an amino acid substitution generated at the putative start codon of copL, caused a loss of copper-inducible transcriptional activation of copA. A nonpolar insertion of a kanamycin resistance gene in copL resulted in copper sensitivity in the wild-type strain. However, repeated attempts to complement copL mutations in trans failed. Analysis of the genomic sequence databases shows that there are copL homologs upstream of copAB genes in X. axonopodis pv. citri, X. campestris pv. campestris, and Xylella fastidiosa. The cloned promoter area upstream of copA in X. axonopodis pv. vesicatoria did not function in Pseudomonas syringae or in E. coli, nor did the P. syringae cop promoter function in Xanthomonas. However, a transcriptional fusion of the Xanthomonas cop promoter with the Pseudomonas copABCDRS was able to confer resistance to copper in Xanthomonas, showing divergence in the mechanisms of regulation of the resistance to copper in phytopathogenic bacteria.

Bacterial Proteins↗
PubMed2002

Identification of a fibronectin-binding protein from Staphylococcus epidermidis.

Staphylococcus epidermidis has been reported to bind to a number of host cell extracellular matrix proteins, including fibronectin. Here we report the identification of a fibronectin-binding protein from S. epidermidis. A phage display library of S. epidermidis genomic DNA was constructed and panned against immobilized fibronectin. A number of phagemid clones containing overlapping inserts were identified, and one of these clones, pSE109FN, contained a 1.4-kb insert. Phage pSE109FN was found to bind to fibronectin but not to collagen, fibrinogen, laminin, or vitronectin. However, pSE109FN also bound to heparin, hyaluronate, and plasminogen, although to a lesser extent than it bound to fibronectin. Analysis of The Institute for Genomic Research S. epidermidis genome sequence database revealed a 1.85-kb region within a putative 30.5-kb open reading frame, to which the overlapping DNA inserts contained within the fibronectin-binding phagemids mapped. We have designated the gene encoding the fibronectin-binding domain embp. A recombinant protein, Embp32, which encompassed the fibronectin-binding domain of Embp, blocked the binding of S. epidermidis, but not the binding of Staphylococcus aureus, to fibronectin. In contrast, a recombinant protein, FnBPB[D1-D4], spanning the fibronectin-binding domain of the S. aureus fibronectin-binding protein FnBPB, blocked binding of S. aureus to fibronectin but had a negligible effect on the binding of S. epidermidis.

Adhesins, Bacterial↗
PubMed2005

Characterization of two novel cryptococcal mannoproteins recognized by immune sera.

Host defenses against the encapsulated yeast Cryptococcus neoformans involve both humoral and cell-mediated immunity. Mannoproteins (MPs) are a heterogeneous class of immunodominant glycoproteins which have been only incompletely characterized. In this study, we report on the molecular features of two novel MPs that are recognized by serum antibodies during cryptococcosis. After fractionation of extracellular cryptococcal products, MPs reacted more strongly than other components with sera from C. neoformans-infected AIDS patients. Further fractionation and Western blot analysis of MPs evidenced the presence of highly reactive bands with molecular masses of 250, 125, 115, and 84 kDa. The 115- and 84-kDa bands contained significant amounts of N-linked oligosaccharides, as shown by decreased molecular mass after peptide-N-glycosidase F treatment. N-terminal amino acid sequences of the two bands were used to search C. neoformans nucleotide databases. Homologous genomic sequences were used to synthesize DNA probes and isolate cDNA clones containing the full-length genes, which were designated MP84 and MP115. Both genes showed the presence of a serine/threonine-rich region, a potential site for heavy glycosylation. MP84 and MP115 showed homology with, respectively, polysaccharide deacetylases and carboxylesterases from other organisms. Recombinant, deglycosylated proteins expressed in Escherichia coli still reacted with sera from patients, albeit more weakly than natural MPs, indicating that at least some of the reactive epitopes were retained in the recombinant forms. In conclusion, we identified two novel MPs that are important targets of antibody responses during cryptococcosis. These data may be useful to devise alternative immunity-based strategies to control the disease.

Amino Acid Sequence↗
PubMed1999

Identification and characterization of SirA, an iron-regulated protein from Staphylococcus aureus.

The acquisition of iron by pathogenic bacteria is often a crucial step in establishing infection. To accomplish this, many bacteria, including Staphylococcus aureus, produce low-molecular-weight iron-chelating siderophores. However, the secretion and transport of these molecules in gram-positive organisms are poorly understood. The sequence, organization, and regulation of genes involved in siderophore transport are conserved among gram-negative bacteria. We used this information to identify a putative siderophore transport locus from an S. aureus genomic sequence database. This locus contains three predicted open reading frames with a high degree of homology to genes involved in siderophore uptake in several bacterial species, in particular the cbr locus of the plant pathogen Erwinia chrysanthemi. The first gene in the locus, which we have designated sir for staphylococcal iron regulated, encodes a putative lipoprotein with a molecular mass of 37 kDa. The open reading frame is preceded by a 19-bp region of dyad symmetry with homology for operator sequences controlling iron-regulated expression of genes in other bacteria. Fur titration experiments indicate that this region of dyad symmetry is sufficient for Fur-dependent regulation in Escherichia coli. The expression of this gene was repressed, in a dose-dependent manner, by the addition of iron to the S. aureus culture medium. sir-encoded proteins may be involved in iron acquisition in vivo and therefore may be targets for antimicrobial agents.

Amino Acid Sequence↗
PubMed2001

The Bordetella bhu locus is required for heme iron utilization.

Bordetella pertussis and Bordetella bronchiseptica are capable of obtaining iron from hemin and hemoglobin. Genes encoding a putative bacterial heme iron acquisition system (bhu, for Bordetella heme utilization) were identified in a B. pertussis genomic sequence database, and the corresponding DNA was isolated from a virulent strain of B. pertussis. A B. pertussis bhuR mutant, predicted to lack the heme outer membrane receptor, was generated by allelic exchange. In contrast to the wild-type strain, bhuR mutant PM5 was incapable of acquiring iron from hemin and hemoglobin; genetic complementation of PM5 with the cloned bhuRSTUV genes restored heme utilization to wild-type levels. In parallel studies, B. bronchiseptica bhu sequences were also identified and a B. bronchiseptica bhuR mutant was constructed and confirmed to be defective in heme iron acquisition. The wild-type B. bronchiseptica parent strain grown under low-iron conditions produced the presumptive BhuR protein, which was absent in the bhuR mutant. Furthermore, production of BhuR by iron-starved B. bronchiseptica was markedly enhanced by culture in hemin-supplemented medium, suggesting that these organisms sense and respond to heme in the environment. Analysis of the genetic region upstream of the bhu cluster identified open reading frames predicted to encode homologs of the Escherichia coli ferric citrate uptake regulators FecI and FecR. These putative Bordetella regulators may mediate heme-responsive positive transcriptional control of the bhu genes.

Amino Acid Sequence↗