Publications

nonlinearity in genetic decoding: homologous dna replicase genes use alternatives of transcriptional slippage or translational frameshifting.	the tau and gamma subunits of dna polymerase iii are both encoded by a single gene in escherichia coli and thermus thermophilus. gamma is two-thirds the size of tau and shares virtually all its amino acid sequence with tau. e. coli and t. thermophilus have evolved very different mechanisms for setting the approximate 1:1 ratio between tau and gamma. both mechanisms put ribosomes into alternate reading frames so that stop codons in the new frame serve to make the smaller gamma protein. in e. coli ...	2000	10677518
use of defined mutants to assess the role of the campylobacter rectus s-layer in bacterium-epithelial cell interactions.	campylobacter rectus is a periodontal pathogen with a 150-kda protein on its cell surface. this protein forms a paracrystalline lattice, called the s-layer, surrounding the outer membrane of this gram-negative bacterium. to initiate a genetic analysis of the possible role of the s-layer in the initial interaction of c. rectus with host epithelial cells, c. rectus strains lacking the s-layer protein gene (crsa) were constructed by allelic exchange mutagenesis. surprisingly, the lack of the s-laye ...	2000	10678961
assembly of archaeal signal recognition particle from recombinant components.	signal recognition particle (srp) takes part in protein targeting and secretion in all organisms. searches for components of archaeal srp in primary databases and completed genomes indicated that archaea possess only homologs of srp rna, and proteins srp19 and srp54. a recombinant srp was assembled from cloned, expressed and purified components of the hyperthermophilic archaeon archaeoglobus fulgidus. recombinant af-srp54 associated with the signal peptide of bovine pre-prolactin translated in v ...	2000	10684931
functionality of purified sigma(n) (sigma(54)) and a nifa-like protein from the hyperthermophile aquifex aeolicus.	the genome sequence of the extremely thermophilic bacterium aquifex aeolicus encodes alternative sigma factor sigma(n) (sigma(54), rpon) and five potential sigma(n)-dependent transcriptional activators. although a. aeolicus possesses no recognizable nitrogenase genes, two of the activators have a high degree of sequence similarity to nifa proteins from nitrogen-fixing proteobacteria. we identified five putative sigma(n)-dependent promoters upstream of operons implicated in functions including su ...	2000	10692367
p(2y) purinoceptor subtypes recruit different mek activators in astrocytes.	extracellular atp can function as a glial trophic factor as well as a neuronal transmitter. in astrocytes, mitogenic signalling by atp is mediated by metabotropic p(2y) receptors that are linked to the extracellular signal regulated protein kinase (erk) cascade, but the types of p(2y) receptors expressed in astrocytes have not been defined and it is not known whether all p(2y) receptor subtypes are coupled to erk by identical or distinct signalling pathways. we found that the p(2y) receptor agon ...	2000	10696092
crystal structure of ribosomal protein l4 shows rna-binding sites for ribosome incorporation and feedback control of the s10 operon.	ribosomal protein l4 resides near the peptidyl transferase center of the bacterial ribosome and may, together with rrna and proteins l2 and l3, actively participate in the catalysis of peptide bond formation. escherichia coli l4 is also an autogenous feedback regulator of transcription and translation of the 11 gene s10 operon. the crystal structure of l4 from thermotoga maritima at 1.7 a resolution shows the protein with an alternating alpha/beta fold and a large disordered loop region. two sep ...	2000	10698923
crystal structure of nad(+)-dependent dna ligase: modular architecture and functional implications.	dna ligases catalyze the crucial step of joining the breaks in duplex dna during dna replication, repair and recombination, utilizing either atp or nad(+) as a cofactor. despite the difference in cofactor specificity and limited overall sequence similarity, the two classes of dna ligase share basically the same catalytic mechanism. in this study, the crystal structure of an nad(+)-dependent dna ligase from thermus filiformis, a 667 residue multidomain protein, has been determined by the multiwav ...	2000	10698952
novel diagnostic algorithm for identification of mycobacteria using genus-specific amplification of the 16s-23s rrna gene spacer and restriction endonucleases.	a novel genus-specific pcr for mycobacteria with simple identification to the species level by restriction fragment length polymorphism (rflp) was established using the 16s-23s ribosomal rna gene (rdna) spacer as a target. panspecificity of primers was demonstrated on the genus level by testing 811 bacterial strains (122 species in 37 genera from 286 reference strains and 525 clinical isolates). all mycobacterial isolates (678 strains among 48 defined species and 5 indeterminate taxons) were amp ...	2000	10699003
aminoacyl-trna synthetases, the genetic code, and the evolutionary process.	the aminoacyl-trna synthetases (aarss) and their relationship to the genetic code are examined from the evolutionary perspective. despite a loose correlation between codon assignments and aars evolutionary relationships, the code is far too highly structured to have been ordered merely through the evolutionary wanderings of these enzymes. nevertheless, the aarss are very informative about the evolutionary process. examination of the phylogenetic trees for each of the aarss reveals the following. ...	2000	10704480
molecular basis of variant pseudo-hurler polydystrophy (mucolipidosis iiic)	mucolipidosis iiic, or variant pseudo-hurler polydystrophy, is an autosomal recessive disease of lysosomal hydrolase trafficking. unlike the related diseases, mucolipidosis ii and iiia, the enzyme affected in mucolipidosis iiic (n-acetylglucosamine-1-phosphotransferase [glcnac-phosphotransferase]) retains full transferase activity on synthetic substrates but lacks activity on lysosomal hydrolases. bovine glcnac-phosphotransferase has recently been isolated as a multisubunit enzyme with the subun ...	2000	10712439
identification of nonpoint sources of fecal pollution in coastal waters by using host-specific 16s ribosomal dna genetic markers from fecal anaerobes.	we describe a new pcr-based method for distinguishing human and cow fecal contamination in coastal waters without culturing indicator organisms, and we show that the method can be used to track bacterial marker sequences in complex environments. we identified two human-specific genetic markers and five cow-specific genetic markers in fecal samples by amplifying 16s ribosomal dna (rdna) fragments from members of the genus bifidobacterium and the bacteroides-prevotella group and performing length ...	2000	10742246
heteroduplex dna and atp induced conformational changes of a muts mismatch repair protein from thermus aquaticus.	atp hydrolysis by muts homologues is required for the function of these proteins in mismatch repair. however, the function of atp hydrolysis in the repair reaction is not very clear. we have examined the role of atp hydrolysis in oligomerization of thermus aquaticus (taq) muts protein in solution. analytical gel filtration and cross-linking of muts protein with disuccinimidyl suburate suggest that taqmuts is a dimer in the presence of atp. atp binding and hydrolysis by taqmuts reduces the hetero ...	2000	10769195
structure of the fmet-trna(fmet)-binding domain of b. stearothermophilus initiation factor if2.	the three-dimensional structure of the fmet-trna(fmet) -binding domain of translation initiation factor if2 from bacillus stearothermophilus has been determined by heteronuclear nmr spectroscopy. its structure consists of six antiparallel beta-strands, connected via loops, and forms a closed beta-barrel similar to domain ii of elongation factors ef-tu and ef-g, despite low sequence homology. two structures of the ternary complexes of the ef-tu small middle dotaminoacyl-trna small middle dot gdp ...	2000	10775275
functional interaction between the werner syndrome protein and dna polymerase delta.	werner syndrome (ws) is an inherited disease characterized by premature onset of aging, increased cancer incidence, and genomic instability. the ws gene encodes a 1,432-amino acid polypeptide (wrn) with a central domain homologous to the recq family of dna helicases. purified wrn unwinds dna with 3'-->5' polarity, and also possesses 3'-->5' exonuclease activity. elucidation of the physiologic function(s) of wrn may be aided by the identification of wrn-interacting proteins. we show here that wrn ...	2000	10781066
a novel amidase (half-amidase) for half-amide hydrolysis involved in the bacterial metabolism of cyclic imides.	a novel amidase involved in bacterial cyclic imide metabolism was purified from blastobacter sp. strain a17p-4. the enzyme physiologically functions in the second step of cyclic imide degradation, i.e., the hydrolysis of monoamidated dicarboxylates (half-amides) to dicarboxylates and ammonia. enzyme production was enhanced by cyclic imides such as succinimide and glutarimide but not by amide compounds which are conventional substrates and inducers of known amidases. the purified amidase showed h ...	2000	10788365
reduction of fe(iii), cr(vi), u(vi), and tc(vii) by deinococcus radiodurans r1.	deinococcus radiodurans is an exceptionally radiation-resistant microorganism capable of surviving acute exposures to ionizing radiation doses of 15,000 gy and previously described as having a strictly aerobic respiratory metabolism. under strict anaerobic conditions, d. radiodurans r1 reduced fe(iii)-nitrilotriacetic acid coupled to the oxidation of lactate to co(2) and acetate but was unable to link this process to growth. d. radiodurans reduced the humic acid analog anthraquinone-2,6-disulfon ...	2000	10788374
contamination and sensitivity issues with a real-time universal 16s rrna pcr.	a set of universal oligonucleotide primers specific for the conserved regions of the eubacterial 16s rrna gene was designed for use with the real-time pcr applied biosystems 7700 (taqman) system. during the development of this pcr, problems were noted with the use of this gene as an amplification target. contamination of reagents with bacterial dna was a major problem exacerbated by the highly sensitive nature of the real-time pcr chemistry. this was compounded by the use of a small amplicon of ...	2000	10790092
rapid identification of yersinia enterocolitica in blood by the 5' nuclease pcr assay.	yersinia enterocolitica accounts for 50% of the clinical sepsis episodes caused by the transfusion of contaminated red blood cells. a 5' nuclease taqman pcr assay was developed to detect y. enterocolitica in blood. primers and a probe based on the nucleotide sequence of the 16s rrna gene from y. enterocolitica were designed. whole-blood samples were spiked with various numbers of y. enterocolitica cells, and total chromosomal dna was extracted. when the taqman pcr assay was performed, as few as ...	2000	10790127
sensitive method for detection of human herpesviruses 6 and 7 in saliva collected in field studies.	to facilitate studies of the epidemiology and natural history of human herpesviruses 6 and 7 in infants, a practical method for collecting and quantifying the dna of these viruses was developed. saliva was collected using small strips of filter paper, and virus was detected using a real-time quantitative fluorescent-probe pcr assay. the sensitivity and specificity of this method even after prolonged drying of the specimens compared favorably to those of our traditional method of collecting and a ...	2000	10790134
the autographa californica nuclear polyhedrosis virus p143 gene encodes a dna helicase.	the p143 protein of autographa californica nuclear polyhedrosis virus is essential for replication of viral dna. to determine the function of p143, the protein was purified to near homogeneity from recombinant baculovirus-infected cells that overexpress p143. atpase activity copurified with p143 protein during purification and also during gel filtration at a high salt concentration. the atpase activity did not require the presence of single-stranded dna, but was stimulated fourfold by the additi ...	2000	10799604
dna polymerase active site is highly mutable: evolutionary consequences.	dna polymerases contain active sites that are structurally superimposable and highly conserved in sequence. to assess the significance of this preservation and to determine the mutational burden that active sites can tolerate, we randomly mutated a stretch of 13 amino acids within the polymerase catalytic site (motif a) of thermus aquaticus dna polymerase i. after selection, by using genetic complementation, we obtained a library of approximately 8, 000 active mutant dna polymerases, of which 35 ...	2000	10805772
listeriolysin o as a reporter to identify constitutive and in vivo-inducible promoters in the pathogen listeria monocytogenes.	listeria monocytogenes is a facultative intracellular gram-positive bacterium capable of growing in the cytoplasm of infected host cells. bacterial escape from the phagosomal vacuole of infected cells is mainly mediated by the pore-forming hemolysin listeriolysin o (llo) encoded by hly. llo-negative mutants of l. monocytogenes are avirulent in the mouse model. we have developed a genetic system with hly as a reporter gene allowing the identification of both constitutive and in vivo-inducible pro ...	2000	10816469
production of exopolysaccharide by lactobacillus rhamnosus r and analysis of its enzymatic degradation during prolonged fermentation.	the potential of lactobacillus rhamnosus r for producing exopolysaccharide (eps) when grown on basal minimum medium supplemented with glucose or lactose was investigated. eps production by l. rhamnosus r is partially growth associated and about 500 mg of eps per liter was synthesized with both sugars. the product yield coefficient (y(eps/s)) was 3.15 (0.0315 g of eps [g of lactose](-1)) and 2.88 (0.0288 g of eps [g of glucose](-1)). it was clearly shown that the amount of eps produced declined u ...	2000	10831403
isolation and expression of lactate dehydrogenase genes from rhizopus oryzae.	rhizopus oryzae is used for industrial production of lactic acid, yet little is known about the genetics of this fungus. in this study i cloned two genes, ldha and ldhb, which code for nad(+)-dependent l-lactate dehydrogenases (ldh) (ec 1.1.1.27), from a lactic acid-producing strain of r. oryzae. these genes are similar to each other and exhibit more than 90% nucleotide sequence identity and they contain no introns. this is the first description of ldh genes in a fungus, and sequence comparisons ...	2000	10831409
use of randomly amplified polymorphic dna as a means of developing genus- and strain-specific streptomyces dna probes.	we have analyzed 20 randomly amplified polymorphic dna (rapd) primers against 36 streptomyces strains, including 17 taxonomically undefined strains, 25 nonstreptomycete actinomycetes, and 12 outgroups consisting of gram-positive and -negative species. most of the primers were useful in identifying unique dna polymorphisms of all strains tested. we have used rapd techniques to develop a genus-specific probe, one not necessarily targeting the ribosomal gene, for streptomyces, and a strain-specific ...	2000	10831438
arabidopsis muts homologs-atmsh2, atmsh3, atmsh6, and a novel atmsh7-form three distinct protein heterodimers with different specificities for mismatched dna.	arabidopsis mismatch repair genes predict muts-like proteins remarkably similar to eukaryotic muts homologs-msh2, msh3, and msh6. a novel feature in arabidopsis is the presence of two msh6-like proteins, designated atmsh6 and atmsh7. combinations of arabidopsis atmsh2 with atmsh3, atmsh6, or atmsh7 proteins-products of in vitro transcription and translation-were analyzed for interactions by analytical gel filtration chromatography. the atmsh2 protein formed heterodimers with atmsh3, atmsh6, and ...	2000	10852942
conservation of sigma-core rna polymerase proximity relationships between the enhancer-independent and enhancer-dependent sigma classes.	two distinct classes of rna polymerase sigma factors (sigma) exist in bacteria and are largely unrelated in primary amino acid sequence and their modes of transcription activation. using tethered iron chelate (fe-babe) derivatives of the enhancer-dependent sigma(54), we mapped several sites of proximity to the beta and beta' subunits of the core rna polymerase. remarkably, most sites localized to those previously identified as close to the enhancer-independent sigma(70) and sigma(38). this indic ...	2000	10856247
male gender predisposes to development of endotoxic shock in the rat.	after intravenous (i.v.) injection of lipopolysaccharide (lps) macrophages release nitric oxide (no) due to the expression of the inducible no synthase (inos). after lps no is abundantly produced also in the cardiovascular system and may contribute to the development of hypotension and shock. since the immune response, the synthesis of no and the regulation of blood pressure (bp) differ between males and females, in the present study the effect of lps on bp, renal function, the plasma and urinar ...	2000	10869545
influence of sulfide and temperature on species composition and community structure of hot spring microbial mats.	in solfataric fields in southwestern iceland, neutral and sulfide-rich hot springs are characterized by thick bacterial mats at 60 to 80 degrees c that are white or yellow from precipitated sulfur (sulfur mats). in low-sulfide hot springs in the same area, grey or pink streamers are formed at 80 to 90 degrees c, and a chloroflexus mat is formed at 65 to 70 degrees c. we have studied the microbial diversity of one sulfur mat (high-sulfide) hot spring and one chloroflexus mat (low-sulfide) hot spr ...	2000	10877776
detection of ralstonia solanacearum strains with a quantitative, multiplex, real-time, fluorogenic pcr (taqman) assay.	a fluorogenic (taqman) pcr assay was developed to detect ralstonia solanacearum strains. two fluorogenic probes were utilized in a multiplex reaction; one broad-range probe (rs) detected all biovars of r. solanacearum, and a second more specific probe (b2) detected only biovar 2a. amplification of the target was measured by the 5' nuclease activity of taq dna polymerase on each probe, resulting in emission of fluorescence. taqman pcr was performed with dna extracted from 42 r. solanacearum and g ...	2000	10877778
a 5' nuclease pcr (taqman) high-throughput assay for detection of the meca gene in staphylococci.	in an effort to find a rapid, efficient, and reliable method of screening large numbers of bacterial isolates for specific antimicrobial resistance genes, we compared conventional pcr results to the results generated using the taqman 5' nuclease pcr kit in conjunction with an abi prism 7700 sequence detector for detecting the meca gene in various species of staphylococci. dna was extracted using two techniques. the first used a high-salt extraction method suitable for conventional pcr but result ...	2000	10878035
detection of human cytomegalovirus dna by real-time quantitative pcr.	a real-time pcr assay was developed to quantify human cytomegalovirus (cmv) dna. this assay was used to demonstrate a higher cmv dna load in plasma of bone marrow transplant patients than in that of blood donors. the cmv load was higher in cmv antigen-positive patients than in antigen-negative patients.	2000	10878073
extensive homologous recombination among widely divergent tt viruses.	analyses of a collection of full-length tt virus genomes showed nearly half of them to be recombinant. the results were highly significant and revealed homologous recombination both within and among genotypes, often involving extremely divergent lineages. recombination breakpoints were significantly more common in the noncoding region of the tt virus genome than in the coding region.	2000	10906223
common fold in helix-hairpin-helix proteins.	helix-hairpin-helix (hhh) is a widespread motif involved in non-sequence-specific dna binding. the majority of hhh motifs function as dna-binding modules, however, some of them are used to mediate protein-protein interactions or have acquired enzymatic activity by incorporating catalytic residues (dna glycosylases). from sequence and structural analysis of hhh-containing proteins we conclude that most hhh motifs are integrated as a part of a five-helical domain, termed (hhh)(2) domain here. it t ...	2000	10908318
a molecular epidemiological study of mycobacterium simiae isolated from aids patients in guadeloupe.	a molecular epidemiological study of mycobacterium simiae strains isolated from aids patients in guadeloupe was performed by the random amplification of polymorphic dna (rapd) and pulsed-field gel electrophoresis (pfge) of drai- or xbai-digested bacterial dnas. a comparison of rapd profiles suggested a similarity of banding patterns within a group of patients (two clusters of two and three patients), but the available epidemiological and clinical information did not support this finding. pfge, o ...	2000	10921982
novel genes coding for lithotrophic sulfur oxidation of paracoccus pantotrophus gb17.	the gene region coding for lithotrophic sulfur oxidation of paracoccus pantotrophus gb17 is located on a 13-kb insert of plasmid peg12. upstream of the previously described six open reading frames (orfs) soxabcdef with a partial sequence of soxa and soxf (c. wodara, f. bardischewsky, and c. g. friedrich, j. bacteriol. 179:5014-5023, 1997), 4,350 bp were sequenced. the sequence completed soxa, and uncovered six new orfs upstream of soxa, designated orf1, orf2, and orf3, and soxxyz. orf1 could enc ...	2000	10940005
reduction of gc --> ta transversion mutation by overexpression of muts in escherichia coli k-12.	overexpression of the muts repair protein significantly decreased the rate of lacz gc --> ta transversion mutation in stationary-phase and exponentially growing bacteria and in muty and mutm mutants, which accumulate mismatches between 8-oxoguanine (8-oxog) and adenine residues in dna. conversely, gc --> ta transversion increased in mutl or muts mutants in stationary phase. in contrast, overexpression of muts did not appreciably reduce lacz at --> cg transversion mutation in a mutt mutant. these ...	2000	10940054
identifying a core rna polymerase surface critical for interactions with a sigma-like specificity factor.	cyclic interactions occurring between a core rna polymerase (rnap) and its initiation factors are critical for transcription initiation, but little is known about subunit interaction. in this work we have identified regions of the single-subunit yeast mitochondrial rnap (rpo41p) important for interaction with its sigma-like specificity factor (mtf1p). previously we found that the whole folded structure of both polypeptides as well as specific amino acids in at least three regions of mtf1p are re ...	2000	10958696
a hydrogen peroxide-forming nadh oxidase that functions as an alkyl hydroperoxide reductase in amphibacillus xylanus.	the amphibacillus xylanus nadh oxidase, which catalyzes the reduction of oxygen to hydrogen peroxide with beta-nadh, can also reduce hydrogen peroxide to water in the presence of free flavin adenine dinucleotide (fad) or the small disulfide-containing salmonella enterica ahpc protein. the enzyme has two disulfide bonds, cys128-cys131 and cys337-cys340, which can act as redox centers in addition to the enzyme-bound fad (k. ohnishi, y. niimura, m. hidaka, h. masaki, h. suzuki, t. uozumi, and t. ni ...	2000	10960086
a positive cis-acting dna element is required for high-level transcription in chlamydia.	the spacer a/t region is a positive cis-acting dna element that was identified in the chlamydia trachomatis rrna promoter region. we have now demonstrated that similar sequences in other chlamydial promoters are important for transcription. substitution of candidate spacer a/t regions in four chlamydial promoters decreased transcription by partially purified c. trachomatis rna polymerase in an in vitro transcription assay. addition of a spacer a/t region to the dnak promoter, which does not cont ...	2000	10960101
application of the 5'-nuclease pcr assay in evaluation and development of methods for quantitative detection of campylobacter jejuni.	campylobacter jejuni is recognized as a leading human food-borne pathogen. traditional diagnostic testing for c. jejuni is not reliable due to special growth requirements and the possibility that this bacterium can enter a viable but nonculturable state. nucleic acid-based tests have emerged as a useful alternative to traditional enrichment testing. in this article, we present a 5'-nuclease pcr assay for quantitative detection of c. jejuni and describe its evaluation. a probe including positions ...	2000	10966425
rapid 5' nuclease (taqman) assay for detection of virulent strains of yersinia enterocolitica.	we have developed a rapid procedure for the detection of virulent yersinia enterocolitica in ground pork by combining a previously described pcr with fluorescent dye technologies. the detection method, known as the fluorogenic 5' nuclease assay (taqman), produces results by measuring the fluorescence produced during pcr amplification, requiring no post-pcr processing. the specificity of the chromosomal yst gene-based assay was tested with 28 bacterial isolates that included 7 pathogenic and 7 no ...	2000	10966441
automated 5' nuclease pcr assay for identification of salmonella enterica.	a simple and ready-to-go test based on a 5' nuclease (taqman) pcr technique was developed for identification of presumptive salmonella enterica isolates. the results were compared with those of conventional methods. the taqman assay was evaluated for its ability to accurately detect 210 s. enterica isolates, including 100 problematic "rough" isolates. an internal positive control was designed to use the same salmonella primers for amplification of a spiked nonrelevant template (116 bp) in the sa ...	2000	10970396
characterization of mselb, a novel mammalian elongation factor for selenoprotein translation.	decoding of uga selenocysteine codons in eubacteria is mediated by the specialized elongation factor selb, which conveys the charged trna(sec) to the a site of the ribosome, through binding to the secis mrna hairpin. in an attempt to isolate the eukaryotic homolog of selb, a database search in this work identified a mouse expressed sequence tag containing the complete cdna encoding a novel protein of 583 amino acids, which we called mselb. several lines of evidence enabled us to establish that m ...	2000	10970870
promoter opening by sigma(54) and sigma(70) rna polymerases: sigma factor-directed alterations in the mechanism and tightness of control.	transcription control at the melting step is not yet understood. here, band shift, cross-linking, and transcription experiments on diverse dna probes were used with two bacterial rna polymerase holoenzymes that differ in how they regulate melting. data indicated that both sigma(54) and sigma(70) holoenzymes assume a default closed form that cannot establish single-strand binding. upon activation the enzymes are converted to an open form that can bind simultaneously to the upstream fork junction ...	2000	10970887
requirement for phe36 for dna binding and mismatch repair by escherichia coli muts protein.	the muts family of dna repair proteins recognizes base pair mismatches and insertion/deletion mismatches and targets them for repair in a strand-specific manner. photocrosslinking and mutational studies previously identified a highly conserved phe residue at the n-terminus of thermus aquaticus muts protein that is critical for mismatch recognition in vitro. here, a mutant escherichia coli muts protein harboring a substitution of ala for the corresponding phe36 residue is assessed for proficiency ...	2000	10982877
dna polymerase i is essential for growth of methylobacterium dichloromethanicum dm4 with dichloromethane.	methylobacterium dichloromethanicum dm4 grows with dichloromethane as the unique carbon and energy source by virtue of a single enzyme, dichloromethane dehalogenase-glutathione s-transferase. a mutant of the dichloromethane-degrading strain m. dichloromethanicum dm4, strain dm4-1445, was obtained by mini-tn5 transposon mutagenesis that was no longer able to grow with dichloromethane. dichloromethane dehalogenase activity in this mutant was comparable to that of the wild-type strain. the site of ...	2000	10986246
uv-induced crosslinks in the 16s rrnas of escherichia coli, bacillus subtilis and thermus aquaticus and their implications for ribosome structure and photochemistry.	sixteen long-range crosslinks are induced in escherichia coli 16s rrna by far-uv irradiation. crosslinking patterns in two other organisms, bacillus subtilis and thermus aquaticus, were investigated to determine if the number and location of crosslinks in e.coli occur because of unusually photoreactive nucleotides at particular locations in the rrna sequence. thirteen long-range crosslinks in b.subtilis and 15 long-range crosslinks in t.aquaticus were detected by gel electrophoresis and 10 cross ...	2000	11000271
application of 5'-nuclease pcr for quantitative detection of listeria monocytogenes in pure cultures, water, skim milk, and unpasteurized whole milk.	pcr techniques have significantly improved the detection and identification of bacterial pathogens. countless adaptations and applications have been described, including quantitative pcr and the latest innovation, real-time pcr. in real-time pcr, e.g., the 5'-nuclease chemistry renders the automated and direct detection and quantification of pcr products possible (p. m. holland et al., proc. natl. acad. sci. usa 88:7276-7280, 1991). we present an assay for the quantitative detection of listeria ...	2000	11010869
eosinophil granule-derived major basic protein induces il-8 expression in human intestinal myofibroblasts.	eosinophil infiltration occurs in a variety of allergic and inflammatory diseases. the release of preformed mediators from eosinophils may contribute to inflammatory responses. we investigated the ability of eosinophil-derived major basic protein and eosinophil-derived neurotoxin to stimulate production of il-8 from intestinal myofibroblasts. intestinal myofibroblasts (18-co cells) were incubated with major basic protein, eosinophil-derived neurotoxin, or a synthetic analogue of major basic prot ...	2000	11012615
mapping the fmet-trna(f)(met) binding site of initiation factor if2.	the interaction between fmet-trna(f)(met) and bacillus stearothermophilus translation initiation factor if2 has been characterized. we demonstrate that essentially all thermodynamic determinants governing the stability and the specificity of this interaction are localized within the acceptor hexanucleotide fmet-3'accaac of the initiator trna and a fairly small area at the surface of the beta-barrel structure of the 90-amino acid c-terminal domain of if2 (if2 c-2). a weak but specific interaction ...	2000	11013225
ribosomal protein l2 is involved in the association of the ribosomal subunits, trna binding to a and p sites and peptidyl transfer.	ribosomal proteins l2, l3 and l4, together with the 23s rna, are the main candidates for catalyzing peptide bond formation on the 50s subunit. that l2 is evolutionarily highly conserved led us to perform a thorough functional analysis with reconstituted 50s particles either lacking l2 or harboring a mutated l2. l2 does not play a dominant role in the assembly of the 50s subunit or in the fixation of the 3'-ends of the trnas at the peptidyl-transferase center. however, it is absolutely required f ...	2000	11013226
pcr performance of the b-type dna polymerase from the thermophilic euryarchaeon thermococcus aggregans improved by mutations in the y-gg/a motif.	the effect of mutations in the highly conserved y-gg/a motif of b-type dna polymerases was studied in the dna polymerase from the hyperthermophilic euryarchaeon thermococcus aggregans. this motif plays a critical role in the balance between the synthesis and degradation of the dna chain. five different mutations of the tyrosine at position 387 (tyr387-->phe, tyr387-->trp, tyr387-->his, tyr387-->asn and tyr387-->ser) revealed that an aromatic ring system is crucial for the synthetic activity of t ...	2000	11024170
rna polymerases from bacillus subtilis and escherichia coli differ in recognition of regulatory signals in vitro.	adaptation of bacterial cells to diverse habitats relies on the ability of rna polymerase to respond to various regulatory signals. some of these signals are conserved throughout evolution, whereas others are species specific. in this study we present a comprehensive comparative analysis of rna polymerases from two distantly related bacterial species, escherichia coli and bacillus subtilis, using a panel of in vitro transcription assays. we found substantial species-specific differences in the a ...	2000	11029421
functional interaction between ssu72 and the rpb2 subunit of rna polymerase ii in saccharomyces cerevisiae.	ssu72 is an essential gene encoding a phylogenetically conserved protein of unknown function that interacts with the general transcription factor tfiib. a recessive ssu72-1 allele was identified as a synthetic enhancer of a tfiib (sua7-1) defect, resulting in a heat-sensitive (ts(-)) phenotype and a dramatic downstream shift in transcription start site selection. here we describe a new allele, ssu72-2, that confers a ts(-) phenotype in a sua7 wild-type background. in an effort to further define ...	2000	11046131
sec-dependent protein export and the involvement of the molecular chaperone secb.		2000	11048650
a new thermoactive pullulanase from desulfurococcus mucosus: cloning, sequencing, purification, and characterization of the recombinant enzyme after expression in bacillus subtilis.	the gene encoding a thermoactive pullulanase from the hyperthermophilic anaerobic archaeon desulfurococcus mucosus (apua) was cloned in escherichia coli and sequenced. apua from d. mucosus showed 45.4% pairwise amino acid identity with the pullulanase from thermococcus aggregans and contained the four regions conserved among all amylolytic enzymes. apua encodes a protein of 686 amino acids with a 28-residue signal peptide and has a predicted mass of 74 kda after signal cleavage. the apua gene wa ...	2000	11053376
development of real-time pcr assays for rapid detection of pfiesteria piscicida and related dinoflagellates.	pfiesteria complex species are heterotrophic and mixotrophic dinoflagellates that have been recognized as harmful algal bloom species associated with adverse fish and human health effects along the east coast of north america, particularly in its largest (chesapeake bay in maryland) and second largest (albermarle-pamlico sound in north carolina) estuaries. in response to impacts on human health and the economy, monitoring programs to detect the organism have been implemented in affected areas. h ...	2000	11055905
pcr bias in ecological analysis: a case study for quantitative taq nuclease assays in analyses of microbial communities.	succession of ecotypes, physiologically diverse strains with negligible rrna sequence divergence, may explain the dominance of small, red-pigmented (phycoerythrin-rich) cyanobacteria in the autotrophic picoplankton of deep lakes (c. postius and a. ernst, arch. microbiol. 172:69-75, 1999). in order to test this hypothesis, it is necessary to determine the abundance of specific ecotypes or genotypes in a mixed background of phylogenetically similar organisms. in this study, we examined the perform ...	2000	11055948
quantitation of pseudomonas aeruginosa in wound biopsy samples: from bacterial culture to rapid 'real-time' polymerase chain reaction.	statement of findings: we developed a real-time detection (rtd) polymerase chain reaction (pcr) with rapid thermal cycling to detect and quantify pseudomonas aeruginosa in wound biopsy samples. this method produced a linear quantitative detection range of 7 logs, with a lower detection limit of 103 colony-forming units (cfu)/g tissue or a few copies per reaction. the time from sample collection to result was less than 1h. rtd-pcr has potential for rapid quantitative detection of pathogens in cri ...	2000	11056755
real-time pcr for quantitative detection of toxoplasma gondii.	the protozoan toxoplasma gondii is one of the most common infectious pathogenic parasites and can cause severe medical complications in infants and immunocompromised individuals. we report here the development of a real-time pcr-based assay for the detection of t. gondii. oligonucleotide primers and a fluorescence-labeled taqman probe were designed to amplify the t. gondii b1 gene. after 40 pcr cycles, the cycle threshold values (c(t)) indicative of the quantity of the target gene were determine ...	2000	11060078
fecal excretion of a novel human circovirus, tt virus, in healthy children.	the role of tt virus (ttv) as a human pathogen is unclear, as is the mode of ttv transmission. to determine the prevalence of ttv infection and the possible fecal-oral route of transmission, we analyzed fecal specimens from 67 healthy, nontransfused children for ttv dna sequences by heminested pcr, using the ng and t primer sets. the overall prevalence of ttv fecal excretion was 22.4% (15 of 67), with the t primer set (19.4%) being more sensitive than the ng primer set (10.4%). ttv prevalence ba ...	2000	11063506
a dual-specificity aminoacyl-trna synthetase in the deep-rooted eukaryote giardia lamblia.	cysteinyl-trna (cys-trna) is essential for protein synthesis. in most organisms the enzyme responsible for the formation of cys-trna is cysteinyl-trna synthetase (cysrs). the only known exceptions are the euryarchaea methanococcus jannaschii and methanobacterium thermoautotrophicum, which do not encode a cysrs. deviating from the accepted concept of one aminoacyl-trna synthetase per amino acid, these organisms employ prolyl-trna synthetase as the enzyme that carries out cys-trna formation. to da ...	2000	11078517
identification of conserved residues contributing to the activities of adenovirus dna polymerase.	adenovirus codes for a dna polymerase that is a member of the dna polymerase alpha family and uses a protein primer for initiation of dna synthesis. it contains motifs characteristic of a proofreading 3'-5'-exonuclease domain located in the n-terminal region and several polymerase motifs located in the c-terminal region. to determine the role of adenovirus dna polymerase in dna replication, 22 site-directed mutations were introduced into the conserved dna polymerase motifs in the c-terminal regi ...	2000	11090167
evidence for horizontal gene transfer in evolution of elongation factor tu in enterococci.	the elongation factor tu, encoded by tuf genes, is a gtp binding protein that plays a central role in protein synthesis. one to three tuf genes per genome are present, depending on the bacterial species. most low-g+c-content gram-positive bacteria carry only one tuf gene. we have designed degenerate pcr primers derived from consensus sequences of the tuf gene to amplify partial tuf sequences from 17 enterococcal species and other phylogenetically related species. the amplified dna fragments were ...	2000	11092850
effects of amplification facilitators on diagnostic pcr in the presence of blood, feces, and meat.	the full potential of diagnostic pcr is limited, in part, by the presence of inhibitors in complex biological samples that reduce the amplification efficiency. therefore, different pre-pcr treatments are being used to reduce the effects of pcr inhibitors. the aim of the present study was to investigate the effects of 16 amplification facilitators to enhance dna amplification in the presence of blood, feces, or meat. different concentrations of amplification facilitators and inhibitory samples we ...	2000	11101581
identification of an outer segment targeting signal in the cooh terminus of rhodopsin using transgenic xenopus laevis.	mislocalization of the photopigment rhodopsin may be involved in the pathology of certain inherited retinal degenerative diseases. here, we have elucidated rhodopsin's targeting signal which is responsible for its polarized distribution to the rod outer segment (ros). various green fluorescent protein (gfp)/rhodopsin cooh-terminal fusion proteins were expressed specifically in the major red rod photoreceptors of transgenic xenopus laevis under the control of the xenopus opsin promoter. the fusio ...	2000	11134067
evolutionary origin, diversification and specialization of eukaryotic muts homolog mismatch repair proteins.	most eubacteria, and all eukaryotes examined thus far, encode homologs of the dna mismatch repair protein muts. although eubacteria encode only one or two muts-like proteins, eukaryotes encode at least six distinct muts homolog (msh) proteins, corresponding to conserved (orthologous) gene families. this suggests evolution of individual gene family lines of descent by several duplication/specialization events. using quantitative phylogenetic analyses (rasa, or relative apparent synapomorphy analy ...	2000	10606644
mechanism of promoter melting by the xeroderma pigmentosum complementation group b helicase of transcription factor iih revealed by protein-dna photo-cross-linking.	the p89/xeroderma pigmentosum complementation group b (xpb) atpase-helicase of transcription factor iih (tfiih) is essential for promoter melting prior to transcription initiation by rna polymerase ii (rnapii). by studying the topological organization of the initiation complex using site-specific protein-dna photo-cross-linking, we have shown that p89/xpb makes promoter contacts both upstream and downstream of the initiation site. the upstream contact, which is in the region where promoter melti ...	2000	11027286
decreased c-src expression enhances osteoblast differentiation and bone formation.	c-src deletion in mice leads to osteopetrosis as a result of reduced bone resorption due to an alteration of the osteoclast. we report that deletion/reduction of src expression enhances osteoblast differentiation and bone formation, contributing to the increase in bone mass. bone histomorphometry showed that bone formation was increased in src null compared with wild-type mice. in vitro, alkaline phosphatase (alp) activity and nodule mineralization were increased in primary calvarial cells and i ...	2000	11038178
endoplasmic reticulum quality control of oligomeric membrane proteins: topogenic determinants involved in the degradation of the unassembled na,k-atpase alpha subunit and in its stabilization by beta subunit assembly.	the molecular nature of determinants that mediate degradation of unassembled, polytopic subunits of oligomeric membrane proteins and their stabilization after partner subunit assembly is largely unknown. expressing truncated na,k-atpase alpha subunits alone or together with beta subunits, we find that in unassembled alpha subunits neither the four n-terminal transmembrane segments acting as efficient alternating signal anchor-stop transfer sequences nor the large, central cytoplasmic loop expose ...	2000	10793142
signal amplification through nucleotide extension and excision on a dendritic dna platform.	techniques that provide strong signal amplification are useful in diagnostic applications, especially in detecting low concentrations of non-amplifiable target molecules. a versatile and strong signal amplification method based on activities of a dna polymerase to generate high concentrations of pyrophosphate (ppi) is described. the generation of ppi is catalyzed by nucleotide extension and excision activities of a dna polymerase on an oligonucleotide cassette. the signal is generated upon enzym ...	2000	10710438
macromolecular mimicry.	some proteins have been shown to mimic the overall shape and structure of nucleic acids. for some of the proteins involved in translating the genetic information into proteins on the ribosome particle, there are indications that such observations of macromolecular mimicry even extend to similarity in interaction with and function on the ribosome. a small number of structural results obtained outside the protein biosynthesis machinery could indicate that the concept of macromolecular mimicry betw ...	2000	10675317
lipopolysaccharide pretreatment protects from renal ischemia/reperfusion injury : possible connection to an interleukin-6-dependent pathway.	in vivo administration of low doses of lipopolysaccharide (lps) to rodents can protect these animals from subsequently administrated, usually lethal doses of endotoxin or lps. in this study we tested the effects of lps pretreatment on ischemia/reperfusion injury in the kidney. male c57/b1 mice were pretreated with different doses of lps or phosphate-buffered saline on days -4 and -3. the right kidney was removed, and the vessels of the left kidney were clamped for 30 or 45 minutes on day 0. crea ...	2000	10623677
search and discovery strategies for biotechnology: the paradigm shift.	profound changes are occurring in the strategies that biotechnology-based industries are deploying in the search for exploitable biology and to discover new products and develop new or improved processes. the advances that have been made in the past decade in areas such as combinatorial chemistry, combinatorial biosynthesis, metabolic pathway engineering, gene shuffling, and directed evolution of proteins have caused some companies to consider withdrawing from natural product screening. in this ...	2000	10974127
escherichia coli rna polymerase core and holoenzyme structures.	multisubunit rna polymerase is an essential enzyme for regulated gene expression. here we report two escherichia coli rna polymerase structures: an 11.0 a structure of the core rna polymerase and a 9.5 a structure of the sigma(70) holoenzyme. both structures were obtained by cryo-electron microscopy and angular reconstitution. core rna polymerase exists in an open conformation. extensive conformational changes occur between the core and the holoenzyme forms of the rna polymerase, which are large ...	2000	11118218
varied molecular interactions at the active sites of several dna polymerases: nonpolar nucleoside isosteres as probes.	we describe a survey of protein-dna interactions with seven different dna polymerases and reverse transcriptases, carried out with nonpolar nucleoside isosteres f (a thymidine analog) and z and q (deoxyadenosine analogues). previous results have shown that z and f can be efficiently replicated opposite each other by the exonuclease-free klenow fragment of dna polymerase i from escherichia coli (kf(-)), although both of them lack watson-crick h-bonding ability. we find that exonuclease-inactive t ...	2000	20882113
rapid, quantitative pcr monitoring of growth of clostridium botulinum type e in modified-atmosphere-packaged fish.	a rapid, quantitative pcr assay (taqman assay) which quantifies clostridium botulinum type e by amplifying a 280-bp sequence from the botulinum neurotoxin type e (bont/e) gene is described. with this method, which uses the hydrolysis of an internal fluoregenic probe and monitors in real time the increase in the intensity of fluorescence during pcr by using the abi prism 7700 sequence detection system, it was possible to perform accurate and reproducible quantification of the c. botulinum type e ...	2001	11133447
the arabidopsis huellenlos gene, which is essential for normal ovule development, encodes a mitochondrial ribosomal protein.	the huellenlos (hll) gene participates in patterning and growth of the arabidopsis ovule. we have isolated the hll gene and shown that it encodes a protein homologous to the l14 proteins of eubacterial ribosomes. the arabidopsis genome also includes a highly similar gene, huellenlos paralog (hlp), and genes for both cytosolic (l23) and chloroplast ribosome l14 proteins. phylogenetic analysis shows that hll and hlp differ significantly from these other two classes of such proteins. hll and hlp fu ...	2001	11752383
directed evolution of polymerase function by compartmentalized self-replication.	we describe compartmentalized self-replication (csr), a strategy for the directed evolution of enzymes, especially polymerases. csr is based on a simple feedback loop consisting of a polymerase that replicates only its own encoding gene. compartmentalization serves to isolate individual self-replication reactions from each other. in such a system, adaptive gains directly (and proportionally) translate into genetic amplification of the encoding gene. csr has applications in the evolution of polym ...	2001	11274352
hyperthermophilic enzymes: sources, uses, and molecular mechanisms for thermostability.	enzymes synthesized by hyperthermophiles (bacteria and archaea with optimal growth temperatures of > 80 degrees c), also called hyperthermophilic enzymes, are typically thermostable (i.e., resistant to irreversible inactivation at high temperatures) and are optimally active at high temperatures. these enzymes share the same catalytic mechanisms with their mesophilic counterparts. when cloned and expressed in mesophilic hosts, hyperthermophilic enzymes usually retain their thermal properties, ind ...	2001	11238984
dna sequences from multiple amplifications reveal artifacts induced by cytosine deamination in ancient dna.	we show that dna molecules amplified by pcr from dna extracted from animal bones and teeth that vary in age between 25 000 and over 50 000 years carry c-->t and g-->a substitutions. these substitutions can reach high proportions among the molecules amplified and are due to the occurrence of modified deoxycytidine residues in the template dna. if the template dna is treated with uracil n-glycosylase, these substitutions are dramatically reduced. they are thus likely to result from deamination of ...	2001	11726688
dna probes using fluorescence resonance energy transfer (fret): designs and applications.	fluorescence resonance energy transfer (fret) is widely used in biomedical research as a reporter method. oligonucleotides with a dna backbone and one or several chromophore tags have found multiple applications as fret probes. they are especially advantageous for the real-time monitoring of biochemical reactions and in vivo studies. this paper reviews the design and applications of various dna-based probes that use fret the approaches used in the design of new dna fret probes are discussed.	2001	11730017
transient dissociation of polyribosomes and concurrent recruitment of calreticulin and calmodulin transcripts in gravistimulated maize pulvini.	the dynamics of polyribosome abundance were studied in gravistimulated maize (zea mays) stem pulvini. during the initial 15 min of gravistimulation, the amount of large polyribosomes transiently decreased. the transient decrease in polyribosome levels was accompanied by a transient decrease in polyribosome-associated mrna. after 30 min of gravistimulation, the levels of polyribosomes and the amount of polyribosome-associated mrna gradually increased over 24 h up to 3- to 4-fold of the initial va ...	2001	11706198
adomet-dependent methylation, dna methyltransferases and base flipping.	twenty adomet-dependent methyltransferases (mtases) have been characterized structurally by x-ray crystallography and nmr. these include seven dna mtases, five rna mtases, four protein mtases and four small molecule mtases acting on the carbon, oxygen or nitrogen atoms of their substrates. the mtases share a common core structure of a mixed seven-stranded beta-sheet (6 downward arrow 7 upward arrow 5 downward arrow 4 downward arrow 1 downward arrow 2 downward arrow 3 downward arrow) referred to ...	2001	11557810
antiapoptotic signaling generated by caspase-induced cleavage of rasgap.	activation of caspases 3 and 9 is thought to commit a cell irreversibly to apoptosis. there are, however, several documented situations (e.g., during erythroblast differentiation) in which caspases are activated and caspase substrates are cleaved with no associated apoptotic response. why the cleavage of caspase substrates leads to cell death in certain cases but not in others is unclear. one possibility is that some caspase substrates generate antiapoptotic signals when cleaved. here we show th ...	2001	11463818
tuning dna "strings": modulating the rate of dna replication with mechanical tension.	recent experiments have measured the rate of replication of dna catalyzed by a single enzyme moving along a stretched template strand. the dependence on tension was interpreted as evidence that t7 and related dna polymerases convert two (n = 2) or more single-stranded template bases to double helix geometry in the polymerization site during each catalytic cycle. however, we find structural data on the t7 enzyme--template complex indicate n = 1. we also present a model for the "tuning" of replica ...	2001	11447284
molecular markers of serine protease evolution.	the evolutionary history of serine proteases can be accounted for by highly conserved amino acids that form crucial structural and chemical elements of the catalytic apparatus. these residues display non- random dichotomies in either amino acid choice or serine codon usage and serve as discrete markers for tracking changes in the active site environment and supporting structures. these markers categorize serine proteases of the chymotrypsin-like, subtilisin-like and alpha/beta-hydrolase fold cla ...	2001	11406580
homogeneous assays for single-nucleotide polymorphism typing using alphascreen.	alphascreen technology allows the development of high-throughput homogeneous proximity assays. in these assays, signal is generated when 680 nm laser light irradiates a donor bead in close proximity to an acceptor bead. for the detection of nucleic acids, donor and acceptor beads are brought into proximity by two bridging probes that hybridize simultaneously to a common target and to the generic oligonucleotides attached covalently to the beads. this method allows the detection of as little as 1 ...	2001	11282975
evaluation of pcr-generated chimeras, mutations, and heteroduplexes with 16s rrna gene-based cloning.	to evaluate pcr-generated artifacts (i.e., chimeras, mutations, and heteroduplexes) with the 16s ribosomal dna (rdna)-based cloning approach, a model community of four species was constructed from alpha, beta, and gamma subdivisions of the division proteobacteria as well as gram-positive bacterium, all of which could be distinguished by hhai restriction digestion patterns. the overall pcr artifacts were significantly different among the three taq dna polymerases examined: 20% for z-taq, with the ...	2001	11157258
function-structure analysis of proteins using covarion-based evolutionary approaches: elongation factors.	the divergent evolution of protein sequences from genomic databases can be analyzed by the use of different mathematical models. the most common treat all sites in a protein sequence as equally variable. more sophisticated models acknowledge the fact that purifying selection generally tolerates variable amounts of amino acid replacement at different positions in a protein sequence. in their "stationary" versions, such models assume that the replacement rate at individual positions remains consta ...	2001	11209054
replicative dna polymerases.	replicative dna polymerases are essential for the replication of the genomes of all living organisms. on the basis of sequence similarities they can be classified into three types. type a polymerases are homologous to bacterial polymerases i, type b comprises archaebacterial dna polymerases and eukaryotic dna polymerase alpha, and the bacterial polymerase iii class make up type c. structures have been solved for several type a and b polymerases, which share a similar architecture. the structure ...	2001	11178285
evaluation of 5' nuclease assay for detection of actinobacillus pleuropneumoniae.	sequence detection by the 5' nuclease taqman assay uses online detection of internal fluorogenic probes in closed pcr tubes. primers and probe were chosen from a part of the omla gene common to all serotypes of actinobacillus pleuropneumoniae, which gave an amplicon of 92 bp. the test was evaluated with 73 lung isolates and 120 tonsil isolates of a. pleuropneumoniae as well as with a collection of reference strains. by using a c(t) value (cycle number in which the fluorescence exceeds the thresh ...	2001	11136780
isolation of segments of homologous genes with only one conserved amino acid region via pcr.	we present a method which allows the isolation of fragments from genes coding for homologous proteins via pcr when only one block of conserved amino acids is available. sets of degenerated primers are defined by reverse translation of the conserved amino acids such that each set contains not more than 128 different sequences. the second primer binding site is provided by a special cassette that is designed such that it does not allow binding of the second primer prior to being copied by dna synt ...	2001	11139638
cloning and sequencing of defective particles derived from the autonomous parvovirus minute virus of mice for the construction of vectors with minimal cis-acting sequences.	the production of wild-type-free stocks of recombinant parvovirus minute virus of mice [mvm(p)] is difficult due to the presence of homologous sequences in vector and helper genomes that cannot easily be eliminated from the overlapping coding sequences. we have therefore cloned and sequenced spontaneously occurring defective particles of mvm(p) with very small genomes to identify the minimal cis-acting sequences required for dna amplification and virus production. one of them has lost all capsid ...	2001	11152501
novel bifunctional hyperthermostable carboxypeptidase/aminoacylase from pyrococcus horikoshii ot3.	genome sequencing of the thermophilic archaeon pyrococcus horikoshii ot3 revealed a gene which had high sequence similarity to the gene encoding the carboxypeptidase of sulfolobus solfataricus and also to that encoding the aminoacylase from bacillus stearothermophilus. the gene from p. horikoshii comprises an open reading frame of 1,164 bp with an atg initiation codon and a tga termination codon, encoding a 43,058-da protein of 387 amino acid residues. however, some of the proposed active-site r ...	2001	11157230
pseudomonas stutzeri nitrite reductase gene abundance in environmental samples measured by real-time pcr.	we used real-time pcr to quantify the denitrifying nitrite reductase gene (nirs), a functional gene of biogeochemical significance. the assay was tested in vitro and applied to environmental samples. the primer-probe set selected was specific for nirs sequences that corresponded approximately to the pseudomonas stutzeri species. the assay was linear from 1 to 10(6) gene copies (r2 = 0.999). variability at low gene concentrations did not allow detection of twofold differences in gene copy number ...	2001	11157241
quantification of human cytomegalovirus dna by real-time pcr.	a quantitative real-time pcr assay was developed to measure human cytomegalovirus (hcmv) dna load in peripheral blood leukocytes (pbls). the hcmv dna load in pbls was normalized by means of the quantification of a cellular gene (albumin). the results of the real-time pcr assay correlated with those of the hcmv pp65-antigenemia assay (p < 0.0001).	2001	11158149
bacterial rna polymerase subunit omega and eukaryotic rna polymerase subunit rpb6 are sequence, structural, and functional homologs and promote rna polymerase assembly.	bacterial dna-dependent rna polymerase (rnap) has subunit composition beta'betaalpha(i)alpha(ii)omega. the role of omega has been unclear. we show that omega is homologous in sequence and structure to rpb6, an essential subunit shared in eukaryotic rnap i, ii, and iii. in escherichia coli, overproduction of omega suppresses the assembly defect caused by substitution of residue 1362 of the largest subunit of rnap, beta'. in yeast, overproduction of rpb6 suppresses the assembly defect caused by th ...	2001	11158566
analysis of escherichia coli strains causing bacteriuria during pregnancy: selection for strains that do not express type 1 fimbriae.	escherichia coli isolates from patients with bacteriuria of pregnancy were compared by pcr with isolates from patients with community-acquired cystitis for the presence of established virulence determinants. the strains from patients with bacteriuria of pregnancy were less likely to carry genes for p-family, s-family, and f1c adhesins, cytotoxic necrotizing factor 1, and aerobactin, but virtually all of the strains carried the genes for type 1 fimbriae. standard mannose-sensitive agglutination o ...	2001	11159970