| pcr-based assay to quantify human immunodeficiency virus type 1 dna in peripheral blood mononuclear cells. | an assay that quantifies the amount of human immunodeficiency virus type 1 (hiv-1) dna in peripheral blood mononuclear cells has been developed. pcr amplification of the hiv-1 dna is performed in the presence of an internal quantitation standard, and colorimetric detection of the amplified product is performed with microwell plates. the copies of hiv-1 dna are normalized to total genomic dna input. the assay has an analytical sensitivity of 10 input copies per amplification reaction and a three- ... | 2000 | 10655358 |
| crystal structure of yeast initiation factor 4a, a dead-box rna helicase. | the eukaryotic translation initiation factor 4a (eif4a) is a member of the dea(d/h)-box rna helicase family, a diverse group of proteins that couples an atpase activity to rna binding and unwinding. previous work has provided the structure of the amino-terminal, atp-binding domain of eif4a. extending those results, we have solved the structure of the carboxyl-terminal domain of eif4a with data to 1.75 a resolution; it has a parallel alpha-beta topology that superimposes, with minor variations, o ... | 2000 | 11087862 |
| 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 |
| recombinant thermus aquaticus rna polymerase, a new tool for structure-based analysis of transcription. | the three-dimensional structure of dna-dependent rna polymerase (rnap) from thermophilic thermus aquaticus has recently been determined at 3.3 a resolution. currently, very little is known about t. aquaticus transcription and no genetic system to study t. aquaticus rnap genes is available. to overcome these limitations, we cloned and overexpressed t. aquaticus rnap genes in escherichia coli. overproduced t. aquaticus rnap subunits assembled into functional rnap in vitro and in vivo when coexpres ... | 2001 | 11114902 |
| protein sequences conserved in prokaryotic aminoacyl-trna synthetases are important for the activity of the processivity factor of human mitochondrial dna polymerase. | previous studies have shown that the small subunit of xenopus dna polymerase gamma (pol gammab) acts as a processivity factor to stimulate the 140 kda catalytic subunit of human dna polymerase gamma. a putative human pol gammab initially identified by analysis of dna sequence had not been shown to be functional, and appeared to be an incomplete clone. in this paper, we report the cloning of full-length human and mouse pol gammab. both human and mouse pol gammab proteins were expressed in their m ... | 2000 | 10666468 |
| cyanobacterial cell walls: news from an unusual prokaryotic envelope. | | 2000 | 10671437 |
| effect of temperature on stability and activity of elongation factor 2 proteins from antarctic and thermophilic methanogens. | despite the presence and abundance of archaea in low-temperature environments, little information is available regarding their physiological and biochemical properties. in order to investigate the adaptation of archaeal proteins to low temperatures, we purified and characterized the elongation factor 2 (ef-2) protein from the antarctic methanogen methanococcoides burtonii, which was expressed in escherichia coli, and compared it to the recombinant ef-2 protein from a phylogenetically related the ... | 2000 | 10671454 |
| vima gene downstream of reca is involved in virulence modulation in porphyromonas gingivalis w83. | a 0.9-kb open reading frame encoding a unique 32-kda protein was identified downstream of the reca gene of porphyromonas gingivalis. reverse transcription-pcr and northern blot analysis showed that both the reca gene and this open reading frame are part of the same transcriptional unit. this cloned fragment was insertionally inactivated using the ermf-ermam antibiotic resistance cassette to create a defective mutant by allelic exchange. when plated on brucella blood agar, the mutant strain, desi ... | 2001 | 11119521 |
| molecular ecology of tetracycline resistance: development and validation of primers for detection of tetracycline resistance genes encoding ribosomal protection proteins. | phylogenetic analysis of tetracycline resistance genes encoding the ribosomal protection proteins (rpps) revealed the monophyletic origin of these genes. the most deeply branching class, exemplified by tet and otra, consisted of genes from the antibiotic-producing organisms streptomyces rimosus and streptomyces lividans. with a high degree of confidence, the corresponding genes of the other seven classes (tet m, tet s, tet o, tet w, tet q, tet t, and tetb p) formed phylogenetically distinct sepa ... | 2001 | 11133424 |
| 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 |
| one tube mutation detection using sensitive fluorescent dyeing of muts protected dna. | a novel, universal method for mutation detection utilising the ability of muts protein to recognise dna incomplementarities is proposed. the examined and reference dna fragments are pcr amplified. the pcr products are purified, mixed, heated and cooled to form heteroduplexes. in the case of mutation the heteroduplex dna containing mismatch is protected against exonuclease digestion by muts, while the dna without mismatches is degraded. the protection effect is visualised by the direct addition o ... | 2000 | 10734213 |
| identification of the outer membrane porin of thermus thermophilus hb8: the channel-forming complex has an unusually high molecular mass and an extremely large single-channel conductance. | the outer membrane of the thermophilic bacterium thermus thermophilus was isolated using sucrose step gradient centrifugation. its detergent extracts contained an ion-permeable channel with an extremely high single-channel conductance of 20 ns in 1 m kcl. the channel protein was purified by preparative sodium dodecyl sulfate (sds)-polyacylamide gel electrophoresis. it has a high molecular mass of 185 kda, and its channel-forming ability resists boiling in sds for 10 min. | 2001 | 11133980 |
| a subunit of human nuclear rnase p has atpase activity. | human nuclear rnase p purified from hela cells has atpase activity. this activity is associated with one of the protein subunits of the enzyme, rpp20. thus, human nuclear rnase p, which contains several proteins and one essential rna, has at least one other enzymatic activity in addition to cleavage of phosphoester bonds in rna. the amino acid sequence of rpp20 has a signature motif found in an atpase-containing subunit of a family of protein complexes (abc transporters) that mediate a variety o ... | 2001 | 11149958 |
| primary structure of a novel subunit in ba3-cytochrome oxidase from thermus thermophilus. | the bax-type cytochrome c oxidase from thermus thermophilus is known as a two subunit enzyme. deduced from the crystal structure of this enzyme, we discovered the presence of an additional transmembrane helix "subunit iia" spanning the membrane. the hydrophobic n-terminally blocked protein was isolated in high yield using high-performance liquid chromatography. its complete amino acid sequence was determined by a combination of automated edman degradation of both the deformylated and the cyanoge ... | 2000 | 11152118 |
| integrity of thermus thermophilus cytochrome c552 synthesized by escherichia coli cells expressing the host-specific cytochrome c maturation genes, ccmabcdefgh: biochemical, spectral, and structural characterization of the recombinant protein. | we describe the design of escherichia coli cells that synthesize a structurally perfect, recombinant cytochrome c from the thermus thermophilus cytochrome c552 gene. key features are (1) construction of a plasmid-borne, chimeric cyca gene encoding an escherichia coli-compatible, n-terminal signal sequence (metlysileseriletyralathrleu alaalaleuserleualaleuproalaglyala) followed by the amino acid sequence of mature thermus cytochrome c552; and (2) coexpression of the chimeric cyca gene with plasmi ... | 2000 | 11152119 |
| a hierarchy of rna subdomains in assembly of the central domain of the 30 s ribosomal subunit. | beginning with the framework that has been developed for the assembly of the 30 s ribosomal subunit, we have identified a series of rnas that are minimal binding sites for proteins s15, s6, s18, and s11 in the central domain from thermus thermophilus. the minimal binding rna for proteins s15, s6, and s18 consists of helix 22 and three-way junctions at both ends composed of portions of helices 20, 21, and 23. addition of the remaining portion of helix 23 to this construct results in the minimal s ... | 2000 | 10744024 |
| development of a reverse transcription-pcr-dna enzyme immunoassay for detection of "norwalk-like" viruses and hepatitis a virus in stool and shellfish. | outbreaks of food- and waterborne gastroenteritis are being increasingly reported throughout the world. the analysis of environmental samples by newer diagnostic techniques such as reverse transcription-pcr (rt-pcr) amplification of nucleic acid has begun to identify human enteric viruses (predominantly "norwalk-like" viruses [nlvs]) as the cause of many of these outbreaks. to streamline nlv detection from environmental samples such as shellfish, we have developed an rt-pcr-oligoprobe amplificat ... | 2001 | 11157239 |
| the crystal structure of the ttcsaa protein: an export-related chaperone from thermus thermophilus. | the csaa protein was first characterized in bacillus subtilis as a molecular chaperone with export-related activities. here we report the 2.0 angstrom-resolution crystal structure of the thermus thermophilus csaa protein, designated ttcsaa. atomic structure and experiments in solution revealed a homodimer as the functional unit. the structure of the ttcsaa monomer is reminiscent of the well known oligonucleotide-binding fold, with the addition of extensions at the n- and c-termini that form an e ... | 2001 | 11157762 |
| structure of the emapii domain of human aminoacyl-trna synthetase complex reveals evolutionary dimer mimicry. | the emapii (endothelial monocyte-activating polypeptide ii) domain is a trna-binding domain associated with several aminoacyl-trna synthetases, which becomes an independent domain with inflammatory cytokine activity upon apoptotic cleavage from the p43 component of the multisynthetase complex. it comprises a domain that is highly homologous to bacterial trna-binding proteins (trbp), followed by an extra domain without homology to known proteins. trbps, which may represent ancient trna chaperones ... | 2001 | 11157763 |
| cloning of the soda gene from corynebacterium melassecola and role of superoxide dismutase in cellular viability. | the soda gene encoding the corynebacterium melassecola manganese-cofactored superoxide dismutase (sod) has been cloned in escherichia coli and sequenced. the gene is transcribed monocistronically; the predicted polypeptide is 200 amino acids long and associates in a homotetrameric, manganese-dependent form, able to complement an sod-deficient e. coli mutant. a second open reading frame, coding for a putative 217-amino-acid protein with high homology to peptide methionine sulfoxide reductases fro ... | 2001 | 11157941 |
| generation of dominant selectable markers for resistance to pseudomonic acid by cloning and mutagenesis of the iles gene from the archaeon methanosarcina barkeri fusaro. | currently, only one selectable marker is available for genetic studies in the archaeal genus methanosarcina. here we report the generation of selectable markers that encode resistance to pseudomonic acid (pa(r)) in methanosarcina species by mutagenesis of the isoleucyl-trna synthetase gene (iles) from methanosarcina barkeri fusaro. the m. barkeri iles gene was obtained by screening of a genomic library for hybridization to a pcr fragment. the complete 3,787-bp dna sequence surrounding and includ ... | 2000 | 10762266 |
| role of the tat ransport system in nitrous oxide reductase translocation and cytochrome cd1 biosynthesis in pseudomonas stutzeri. | by transforming n2o to n2, the multicopper enzyme nitrous oxide reductase provides a periplasmic electron sink for a respiratory chain that is part of denitrification. the signal sequence of the enzyme carries the heptameric twin-arginine consensus motif characteristic of the tat pathway. we have identified tat genes of pseudomonas stutzeri and functionally analyzed the unlinked tatc and tate loci. a tatc mutant retained n2o reductase in the cytoplasm in the unprocessed form and lacking the meta ... | 2001 | 11160097 |
| structure and mechanism of the aberrant ba(3)-cytochrome c oxidase from thermus thermophilus. | cytochrome c oxidase is a respiratory enzyme catalysing the energy-conserving reduction of molecular oxygen to water. the crystal structure of the ba(3)-cytochrome c oxidase from thermus thermophilus has been determined to 2.4 a resolution using multiple anomalous dispersion (mad) phasing and led to the discovery of a novel subunit iia. a structure-based sequence alignment of this phylogenetically very distant oxidase with the other structurally known cytochrome oxidases leads to the identificat ... | 2000 | 10775261 |
| kh domain: one motif, two folds. | the k homology (kh) module is a widespread rna-binding motif that has been detected by sequence similarity searches in such proteins as heterogeneous nuclear ribonucleoprotein k (hnrnp k) and ribosomal protein s3. analysis of spatial structures of kh domains in hnrnp k and s3 reveals that they are topologically dissimilar and thus belong to different protein folds. thus kh motif proteins provide a rare example of protein domains that share significant sequence similarity in the motif regions but ... | 2001 | 11160884 |
| characterization of rnase p from thermotoga maritima. | the protein subunit of rnase p from a thermophilic bacterium, thermotoga maritima, was overexpressed in and purified from escherichia coli. the cloned protein was reconstituted with the rna subunit transcribed in vitro. the temperature optimum of the holoenzyme is near 50 degrees c, with no enzymatic activity at 65 degrees c or above. this finding is in sharp contrast to the optimal growth temperature of t.maritima, which is near 80 degrees c. however, in heterologous reconstitution experiments ... | 2001 | 11160919 |
| crystal structure of the holliday junction migration motor protein ruvb from thermus thermophilus hb8. | we report here the crystal structure of the ruvb motor protein from thermus thermophilus hb8, which drives branch migration of the holliday junction during homologous recombination. ruvb has a crescent-like architecture consisting of three consecutive domains, the first two of which are involved in atp binding and hydrolysis. dna is likely to interact with a large basic cleft, which encompasses the atp-binding pocket and domain boundaries, whereas the junction-recognition protein ruva may bind a ... | 2001 | 11171970 |
| connection between poly-beta-hydroxybutyrate biosynthesis and growth on c(1) and c(2) compounds in the methylotroph methylobacterium extorquens am1. | several dna regions containing genes involved in poly-beta-hydroxybutyrate (phb) biosynthesis and degradation and also in fatty acid degradation were identified from genomic sequence data and have been characterized in the serine cycle facultative methylotroph methylobacterium extorquens am1. genes involved in phb biosynthesis include those encoding beta-ketothiolase (phaa), nadph-linked acetoacetyl coenzyme a (acetyl-coa) reductase (phab), and phb synthase (phac). phaa and phab are closely link ... | 2001 | 11208803 |
| experimental evolution of enzyme temperature activity profile: selection in vivo and characterization of low-temperature-adapted mutants of pyrococcus furiosus ornithine carbamoyltransferase. | we have obtained mutants of pyrococcus furiosus ornithine carbamoyltransferase active at low temperatures by selecting for complementation of an appropriate yeast mutant after in vivo mutagenesis. the mutants were double ones, still complementing at 15 degrees c, a temperature already in the psychrophilic range. their kinetic analysis is reported. | 2001 | 11208811 |
| occurrence of transsulfuration in synthesis of l-homocysteine in an extremely thermophilic bacterium, thermus thermophilus hb8. | a cell extract of an extremely thermophilic bacterium, thermus thermophilus hb8, cultured in a synthetic medium catalyzed cystathionine gamma-synthesis with o-acetyl-l-homoserine and l-cysteine as substrates but not beta-synthesis with dl-homocysteine and l-serine (or o-acetyl-l-serine). the amounts of synthesized enzymes metabolizing sulfur-containing amino acids were estimated by determining their catalytic activities in cell extracts. the syntheses of cystathionine beta-lyase (ec 4.4.1.8) and ... | 2001 | 11222609 |
| detection of infectious cryptosporidium parvum oocysts in mussels (mytilus galloprovincialis) and cockles (cerastoderma edule). | infective cryptosporidium parvum oocysts were detected in mussels (mytilus galloprovincialis) and cockles (cerastoderma edule) from a shellfish-producing region (gallaecia, northwest spain, bounded by the atlantic ocean) that accounts for the majority of european shellfish production. shellfish were collected from bay sites with different degrees of organic pollution. shellfish harboring c. parvum oocysts were recovered only from areas located near the mouths of rivers with a high density of gra ... | 2000 | 10788352 |
| genome of the extremely radiation-resistant bacterium deinococcus radiodurans viewed from the perspective of comparative genomics. | the bacterium deinococcus radiodurans shows remarkable resistance to a range of damage caused by ionizing radiation, desiccation, uv radiation, oxidizing agents, and electrophilic mutagens. d. radiodurans is best known for its extreme resistance to ionizing radiation; not only can it grow continuously in the presence of chronic radiation (6 kilorads/h), but also it can survive acute exposures to gamma radiation exceeding 1,500 kilorads without dying or undergoing induced mutation. these characte ... | 2001 | 11238985 |
| crystal structure of hiv-1 reverse transcriptase in complex with a polypurine tract rna:dna. | we have determined the 3.0 a resolution structure of wild-type hiv-1 reverse transcriptase in complex with an rna:dna oligonucleotide whose sequence includes a purine-rich segment from the hiv-1 genome called the polypurine tract (ppt). the ppt is resistant to ribonuclease h (rnase h) cleavage and is used as a primer for second dna strand synthesis. the 'rnase h primer grip', consisting of amino acids that interact with the dna primer strand, may contribute to rnase h catalysis and cleavage spec ... | 2001 | 11250910 |
| prediction of structural domains of tap reveals details of its interaction with p15 and nucleoporins. | vertebrate tap is a nuclear mrna export factor homologous to yeast mex67p. the middle domain of tap binds directly to p15, a protein related to the nuclear transport factor 2 (ntf2), whereas its c-terminal domain interacts with various nucleoporins, the components of the nuclear pore complex (npc). here, we report that the middle domain of tap is also similar to ntf2, as well as to regions in ras-gap sh3 domain binding protein (g3bp) and some plant protein kinases. based on the known three-dimen ... | 2000 | 11256625 |
| peptide deformylase as an antibacterial drug target: assays for detection of its inhibition in escherichia coli cell homogenates and intact cells. | an assay was developed to determine the activity of peptide deformylase (pdf) inhibitors under conditions as close as possible to the physiological situation. the assay principle is the detection of n-terminal [35s]methionine labeling of a protein that contains no internal methionine. if pdf is active, the deformylation of the methionine renders the peptide a substrate for methionine aminopeptidase, resulting in the removal of the n-terminal methionine label. in the presence of a pdf inhibitor, ... | 2001 | 11257015 |
| peptide deformylase as an antibacterial drug target: target validation and resistance development. | new inhibitors of peptide deformylase (pdf) which are very potent against the isolated enzyme and show a certain degree of antibacterial activity have recently been synthesized by our group. several lines of experimental evidence indicate that these inhibitors indeed interfere with the target enzyme in the bacterial cell. (i) the inhibition of escherichia coli growth could be counteracted by overexpression of pdf from different organisms, including e. coli, streptococcus pneumoniae, and haemophi ... | 2001 | 11257016 |
| transcriptional regulation of the cpr gene cluster in ortho-chlorophenol-respiring desulfitobacterium dehalogenans. | to characterize the expression and possible regulation of reductive dehalogenation in halorespiring bacteria, a 11.5-kb genomic fragment containing the o-chlorophenol reductive dehalogenase-encoding cprba genes of the gram-positive bacterium desulfitobacterium dehalogenans was subjected to detailed molecular characterization. sequence analysis revealed the presence of eight designated genes with the order cprtkzebacd and with the same polarity except for cprt. the deduced cprc and cprk gene prod ... | 2000 | 11004165 |
| rac, a stable ribosome-associated complex in yeast formed by the dnak-dnaj homologs ssz1p and zuotin. | the yeast cytosol contains multiple homologs of the dnak and dnaj chaperone family. our current understanding of which homologs functionally interact is incomplete. zuotin is a dnaj homolog bound to the yeast ribosome. we have now identified the dnak homolog ssz1p/pdr13p as zuotin's partner chaperone. zuotin and ssz1p form a ribosome-associated complex (rac) that is bound to the ribosome via the zuotin subunit. rac is unique among the eukaryotic dnak-dnaj systems, as the 1:1 complex is stable, e ... | 2001 | 11274393 |
| bacterial diversity and community structure in an aerated lagoon revealed by ribosomal intergenic spacer analyses and 16s ribosomal dna sequencing. | we investigated the bacterial community structure in an aerated plug-flow lagoon treating pulp and paper mill effluent. for this investigation, we developed a composite method based on analyses of pcr amplicons containing the ribosomal intergenic spacer (ris) and its flanking partial 16s rrna gene. community percent similarity was determined on the basis of ris length polymorphism. a community succession was evident in the lagoon, indicated by a progressive community transition through seven sam ... | 2001 | 11282606 |
| purification and characterization of the recombinant thermus sp. strain t2 alpha-galactosidase expressed in escherichia coli. | the nucleotide sequence of the thermus sp. strain t2 dna coding for a thermostable alpha-galactosidase was determined. the deduced amino acid sequence of the enzyme predicts a polypeptide of 474 amino acids (m(r), 53,514). the observed homology between the deduced amino acid sequences of the enzyme and alpha-galactosidase from thermus brockianus was over 70%. thermus sp. strain t2 alpha-galactosidase was expressed in its active form in escherichia coli and purified. native polyacrylamide gel ele ... | 2001 | 11282611 |
| elongation of repetitive dna by dna polymerase from a hyperthermophilic bacterium thermus thermophilus. | short repetitive dna sequences are believed to be one of the primordial genetic elements that served as a source of complex large dna found in the genome of modern organisms. however, the mechanism of its expansion (increase in repeat number) during the course of evolution is unclear. we demonstrate that the dna polymerase of the hyperthermophilic bacterium thermus thermophilus can elongate oligodna with several tandem repeats to very long dna in vitro. for instance, 48mer repetitive oligodna (t ... | 2000 | 11024180 |
| rna tertiary interactions in the large ribosomal subunit: the a-minor motif. | analysis of the 2.4-a resolution crystal structure of the large ribosomal subunit from haloarcula marismortui reveals the existence of an abundant and ubiquitous structural motif that stabilizes rna tertiary and quaternary structures. this motif is termed the a-minor motif, because it involves the insertion of the smooth, minor groove edges of adenines into the minor groove of neighboring helices, preferentially at c-g base pairs, where they form hydrogen bonds with one or both of the 2' ohs of ... | 2001 | 11296253 |
| two-dimensional gel electrophoresis analyses of ph-dependent protein expression in facultatively alkaliphilic bacillus pseudofirmus of4 lead to characterization of an s-layer protein with a role in alkaliphily. | the large majority of proteins of alkaliphilic bacillus pseudofirmus of4 grown at ph 7.5 and 10.5, as studied by two-dimensional gel electrophoresis analyses, did not exhibit significant ph-dependent variation. a new surface layer protein (slpa) was identified in these studies. although the prominence of some apparent breakdown products of slpa in gels from ph 10.5-grown cells led to discovery of the alkaliphile s-layer, the largest and major slpa forms were present in large amounts in gels from ... | 2000 | 11029415 |
| anaerobic xylose fermentation by recombinant saccharomyces cerevisiae carrying xyl1, xyl2, and xks1 in mineral medium chemostat cultures. | for ethanol production from lignocellulose, the fermentation of xylose is an economic necessity. saccharomyces cerevisiae has been metabolically engineered with a xylose-utilizing pathway. however, the high ethanol yield and productivity seen with glucose have not yet been achieved. to quantitatively analyze metabolic fluxes in recombinant s. cerevisiae during metabolism of xylose-glucose mixtures, we constructed a stable xylose-utilizing recombinant strain, tmb 3001. the xyl1 and xyl2 genes fro ... | 2000 | 10919795 |
| cloning and functional characterization of an nad(+)-dependent dna ligase from staphylococcus aureus. | a staphylococcus aureus mutant conditionally defective in dna ligase was identified by isolation of complementing plasmid clones that encode the s. aureus liga gene. orthologues of the putative s. aureus nad(+)-dependent dna ligase could be identified in the genomes of bacillus stearothermophilus and other gram-positive bacteria and confirmed the presence of four conserved amino acid motifs, including motif i, kxdg with lysine 112, which is believed to be the proposed site of adenylation. dna se ... | 2001 | 11325928 |
| evidence against an interaction between the mrna downstream box and 16s rrna in translation initiation. | based on the complementarity of the initial coding region (downstream box [db]) of several bacterial and phage mrnas to bases 1469 to 1483 in helix 44 of 16s rrna (anti-downstream box [adb]), it has been proposed that db-adb base pairing enhances translation in a way that is similar to that of the shine-dalgarno (sd)/anti-shine-dalgarno (asd) interaction. computer modeling of helix 44 on the 30s subunit shows that the topography of the 30s ribosome does not allow a simultaneous db-adb interactio ... | 2001 | 11344158 |
| crystal structure of a repair enzyme of oxidatively damaged dna, mutm (fpg), from an extreme thermophile, thermus thermophilus hb8. | the mutm [formamidopyrimidine dna glycosylase (fpg)] protein is a trifunctional dna base excision repair enzyme that removes a wide range of oxidatively damaged bases (n-glycosylase activity) and cleaves both the 3'- and 5'-phosphodiester bonds of the resulting apurinic/apyrimidinic site (ap lyase activity). the crystal structure of mutm from an extreme thermophile, thermus thermophilus hb8, was determined at 1.9 a resolution with multiwavelength anomalous diffraction phasing using the intrinsic ... | 2000 | 10921868 |
| specific interaction between the ribosome recycling factor and the elongation factor g from mycobacterium tuberculosis mediates peptidyl-trna release and ribosome recycling in escherichia coli. | once the translating ribosomes reach a termination codon, the nascent polypeptide chain is released in a factor-dependent manner. however, the p-site-bound deacylated trna and the ribosomes themselves remain bound to the mrna (post-termination complex). the ribosome recycling factor (rrf) plays a vital role in dissociating this complex. here we show that the mycobacterium tuberculosis rrf (mturrf) fails to rescue escherichia coli lj14, a strain temperature-sensitive for rrf (frr(ts)). more inter ... | 2001 | 11387230 |
| visualization of trna movements on the escherichia coli 70s ribosome during the elongation cycle. | three-dimensional cryomaps have been reconstructed for trna-ribosome complexes in pre- and posttranslocational states at 17-a resolution. the positions of trnas in the a and p sites in the pretranslocational complexes and in the p and e sites in the posttranslocational complexes have been determined. of these, the p-site trna position is the same as seen earlier in the initiation-like fmet-trna(f)(met)-ribosome complex, where it was visualized with high accuracy. now, the positions of the a- and ... | 2000 | 10931859 |
| using surface-bound rubidium ions for protein phasing. | rubidium is a monovalent metal that can be used as a counterion in protein solutions. x-ray anomalous scattering from rubidium ions bound to the protein surface was used for phasing of the crystal structure of the hsp60 apical domain from thermus thermophilus. multiple-wavelength anomalous dispersion (mad) data were collected from a crystal obtained from a solution containing 0.2 m rubidium salt. one molecule of protein (147 amino acids) binds one well ordered and one poorly ordered rb atom. pha ... | 2001 | 11418770 |
| characterization of a heme-dependent catalase from methanobrevibacter arboriphilus. | recently it was reported that methanogens of the genus methanobrevibacter exhibit catalase activity. this was surprising, since methanobrevibacter species belong to the order methanobacteriales, which are known not to contain cytochromes and to lack the ability to synthesize heme. we report here that methanobrevibacter arboriphilus strains az and dh1 contained catalase activity only when the growth medium was supplemented with hemin. the heme catalase was purified and characterized, and the enco ... | 2001 | 11425719 |
| natural transformation in mesophilic and thermophilic bacteria: identification and characterization of novel, closely related competence genes in acinetobacter sp. strain bd413 and thermus thermophilus hb27. | the mesophile acinetobacter sp. strain bd413 and the extreme thermophile thermus thermophilus hb27 display high frequencies of natural transformation. in this study we identified and characterized a novel competence gene in acinetobacter sp. strain bd413, coma, whose product displays significant similarities to the competence proteins coma and comec in neisseria and bacillus species. transcription of coma correlated with growth phase-dependent transcriptional regulation of the recently identifie ... | 2001 | 11425734 |
| molecular characterization of desulfovibrio gigas neelaredoxin, a protein involved in oxygen detoxification in anaerobes. | desulfovibrio gigas neelaredoxin is an iron-containing protein of 15 kda, having a single iron site with a his(4)cys coordination. neelaredoxins and homologous proteins are widespread in anaerobic prokaryotes and have superoxide-scavenging activity. to further understand its role in anaerobes, its genomic organization and expression in d. gigas were studied and its ability to complement escherichia coli superoxide dismutase deletion mutant was assessed. in d. gigas, neelaredoxin is transcribed a ... | 2001 | 11443075 |
| tarp: a nuclear protein expressed in prostate and breast cancer cells derived from an alternate reading frame of the t cell receptor gamma chain locus. | previously, we identified the expression of a prostate-specific form of t cell receptor gamma chain (tcrgamma) mrna in the human prostate and demonstrated that it originates from epithelial cells and not from infiltrating t lymphocytes. here, we show that this prostate-specific transcript is also expressed in three breast cancer cell lines and breast cancer tissues. analysis of the cdna sequence predicts that this transcript can encode two protein products of 7 and 13 kda, and in vitro translati ... | 2000 | 10931945 |
| high stability of a ferredoxin from the hyperthermophilic archaeon a. ambivalens: involvement of electrostatic interactions and cofactors. | the ferredoxin from the thermophilic archaeon acidianus ambivalens is a small monomeric seven-iron protein with a thermal midpoint (t(m)) of 122 degrees c (ph 7). to gain insight into the basis of its thermostability, we have characterized unfolding reactions induced chemically and thermally at various phs. thermal unfolding of this ferredoxin, in the presence of various guanidine hydrochloride (guhcl) concentrations, yields a linear correlation between unfolding enthalpies (deltah[t(m)]) and t( ... | 2001 | 11468351 |
| assessment, by transcription-mediated amplification, of virologic response in patients with chronic hepatitis c virus treated with peginterferon alpha-2a. | transcription-mediated amplification (tma) is an isothermal, autocatalytic target amplification method which has the potential to detect less than 50 hepatitis c virus (hcv) rna copies/ml (10 iu/ml). the tma assay was used to assess the presence of residual hcv rna in plasma from patients treated with polyethylene glycol-modified interferon alpha-2a (peginterferon alpha-2a) who showed a virologic relapse after the end of therapy. stored end-of-treatment and end-of-follow-up plasma samples from 1 ... | 2001 | 11474002 |
| the kink-turn: a new rna secondary structure motif. | analysis of the haloarcula marismortui large ribosomal subunit has revealed a common rna structure that we call the kink-turn, or k-turn. the six k-turns in h.marismortui 23s rrna superimpose with an r.m.s.d. of 1.7 a. there are two k-turns in the structure of thermus thermophilus 16s rrna, and the structures of u4 snrna and l30e mrna fragments form k-turns. the structure has a kink in the phosphodiester backbone that causes a sharp turn in the rna helix. its asymmetric internal loop is flanked ... | 2001 | 11483524 |
| genetic analysis of an incomplete muts gene from pseudomonas putida. | we genetically characterized the pseudomonas putida muts gene and found that it encodes a smaller muts protein than do the genes of other bacteria. this gene is able to function in the muts mutants of escherichia coli and bacillus subtilis. a p. putida muts mutant has a mutation frequency 1,000-fold greater than that of the wild-type strain. | 2000 | 10960118 |
| thermostable chitosanase from bacillus sp. strain ck4: cloning and expression of the gene and characterization of the enzyme. | a thermostable chitosanase gene from the environmental isolate bacillus sp. strain ck4, which was identified on the basis of phylogenetic analysis of the 16s rrna gene sequence and phenotypic analysis, was cloned, and its complete dna sequence was determined. the thermostable chitosanase gene was composed of an 822-bp open reading frame which encodes a protein of 242 amino acids and a signal peptide corresponding to a 30-kda enzyme. the deduced amino acid sequence of the chitosanase from bacillu ... | 2000 | 10966383 |
| structure and activity of clpb from escherichia coli. role of the amino-and -carboxyl-terminal domains. | clpb is a member of a protein-disaggregating multi-chaperone system in escherichia coli. the mechanism of protein-folding reactions mediated by clpb is currently unknown, and the functional role of different sequence regions in clpb is under discussion. we have expressed and purified the full-length clpb and three truncated variants with the n-terminal, c-terminal, and a double n- and c-terminal deletion. we studied the protein concentration-dependent and atp-induced oligomerization of clpb, cas ... | 2000 | 10982797 |
| high-throughput genotyping of single nucleotide polymorphisms with rolling circle amplification. | single nucleotide polymorphisms (snps) are the foundation of powerful complex trait and pharmacogenomic analyses. the availability of large snp databases, however, has emphasized a need for inexpensive snp genotyping methods of commensurate simplicity, robustness, and scalability. we describe a solution-based, microtiter plate method for snp genotyping of human genomic dna. the method is based upon allele discrimination by ligation of open circle probes followed by rolling circle amplification o ... | 2001 | 11511324 |
| a dna ligase from a hyperthermophilic archaeon with unique cofactor specificity. | a gene encoding dna ligase (lig(tk)) from a hyperthermophilic archaeon, thermococcus kodakaraensis kod1, has been cloned and sequenced, and its protein product has been characterized. lig(tk) consists of 1,686 bp, corresponding to a polypeptide of 562 amino acids with a predicted molecular mass of 64,079 da. sequence comparison with previously reported dna ligases and the presence of conserved motifs suggested that lig(tk) was an atp-dependent dna ligase. phylogenetic analysis indicated that lig ... | 2000 | 11053387 |
| 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 |
| a conformational change in the ribosomal peptidyl transferase center upon active/inactive transition. | the ribosome is a dynamic particle that undergoes many structural changes during translation. we show through chemical probing with dimethyl sulfate (dms) that conformational changes occur at several nucleotides in the peptidyl transferase center upon alterations in ph, temperature, and monovalent ion concentration, consistent with observations made by elson and coworkers over 30 years ago. moreover, we have found that the ph-dependent dms reactivity of a2451 in the center of the 23s rrna peptid ... | 2001 | 11517305 |
| production of recombinant alpha-galactosidases in thermus thermophilus. | a thermus thermophilus selector strain for production of thermostable and thermoactive alpha-galactosidase was constructed. for this purpose, the native alpha-galactosidase gene (agat) of t. thermophilus th125 was inactivated to prevent background activity. in our first attempt, insertional mutagenesis of agat by using a cassette carrying a kanamycin resistance gene led to bacterial inability to utilize melibiose (alpha-galactoside) and galactose as sole carbohydrate sources due to a polar effec ... | 2001 | 11526023 |
| xylulokinase overexpression in two strains of saccharomyces cerevisiae also expressing xylose reductase and xylitol dehydrogenase and its effect on fermentation of xylose and lignocellulosic hydrolysate. | fermentation of the pentose sugar xylose to ethanol in lignocellulosic biomass would make bioethanol production economically more competitive. saccharomyces cerevisiae, an efficient ethanol producer, can utilize xylose only when expressing the heterologous genes xyl1 (xylose reductase) and xyl2 (xylitol dehydrogenase). xylose reductase and xylitol dehydrogenase convert xylose to its isomer xylulose. the gene xks1 encodes the xylulose-phosphorylating enzyme xylulokinase. in this study, we determi ... | 2001 | 11526030 |
| the structure of an asprs-trna(asp) complex reveals a trna-dependent control mechanism. | the 2.6 a resolution crystal structure of an inactive complex between yeast trna(asp) and escherichia coli aspartyl-trna synthetase reveals the molecular details of a trna-induced mechanism that controls the specificity of the reaction. the dimer is asymmetric, with only one of the two bound trnas entering the active site cleft of its subunit. however, the flipping loop, which controls the proper positioning of the amino acid substrate, acts as a lid and prevents the correct positioning of the t ... | 2001 | 11566892 |
| multiple lateral transfers of dissimilatory sulfite reductase genes between major lineages of sulfate-reducing prokaryotes. | a large fragment of the dissimilatory sulfite reductase genes (dsrab) was pcr amplified and fully sequenced from 30 reference strains representing all recognized lineages of sulfate-reducing bacteria. in addition, the sequence of the dsrab gene homologs of the sulfite reducer desulfitobacterium dehalogenans was determined. in contrast to previous reports, comparative analysis of all available dsrab sequences produced a tree topology partially inconsistent with the corresponding 16s rrna phylogen ... | 2001 | 11567003 |
| characterization of a highly thermostable alkaline phosphatase from the euryarchaeon pyrococcus abyssi. | this work reports the first isolation and characterization of an alkaline phosphatase (ap) from a hyperthermophilic archaeon. an ap gene from pyrococcus abyssi, a euryarchaeon isolated from a deep-sea hydrothermal vent, was cloned and the enzyme expressed in escherichia coli. analysis of the sequence showed conservation of the active site and structural elements of the e. coli ap. the recombinant ap was purified and characterized. monomeric and homodimeric active forms were detected, with a mono ... | 2001 | 11571149 |
| mutations in the listerial prob gene leading to proline overproduction: effects on salt tolerance and murine infection. | the observed sensitivity of listeria monocytogenes to the toxic proline analogue l-azetidine-2-carboxylic acid (az) suggested that proline synthesis in listeria may be regulated by feedback inhibition of gamma-glutamyl kinase (gk), the first enzyme of the proline biosynthesis pathway, encoded by the prob gene. taking advantage of the epicurian coli mutator strain xl1-red, we performed random mutagenesis of the recently described proba operon and generated three independent mutations in the liste ... | 2001 | 11571156 |
| screening of active lyssavirus infection in wild bat populations by viral rna detection on oropharyngeal swabs. | brain analysis cannot be used for the investigation of active lyssavirus infection in healthy bats because most bat species are protected by conservation directives. consequently, serology remains the only tool for performing virological studies on natural bat populations; however, the presence of antibodies merely reflects past exposure to the virus and is not a valid marker of active infection. this work describes a new nested reverse transcription (rt)-pcr technique specifically designed for ... | 2001 | 11574590 |
| visualization of protein s1 within the 30s ribosomal subunit and its interaction with messenger rna. | s1 is the largest ribosomal protein, present in the small subunit of the bacterial ribosome. it has a pivotal role in stabilizing the mrna on the ribosome. thus far, s1 has eluded structural determination. we have identified the s1 protein mass in the cryo-electron microscopic map of the escherichia coli ribosome by comparing the map with a recent x-ray crystallographic structure of the 30s subunit, which lacks s1. according to our finding, s1 is located at the junction of head, platform, and ma ... | 2001 | 11593008 |
| characterization of a brucella species 25-kilobase dna fragment deleted from brucella abortus reveals a large gene cluster related to the synthesis of a polysaccharide. | in the present study we completed the nucleotide sequence of a brucella melitensis 16m dna fragment deleted from b. abortus that accounts for 25,064 bp and show that the other brucella spp. contain the entire 25-kb dna fragment. two short direct repeats of four nucleotides, detected in the b. melitensis 16m dna flanking both sides of the fragment deleted from b. abortus, might have been involved in the deletion formation by a strand slippage mechanism during replication. in addition to omp31, co ... | 2001 | 11598046 |
| sequence analysis of four shigella boydii o-antigen loci: implication for escherichia coli and shigella relationships. | shigella strains are in reality clones of escherichia coli and are believed to have emerged relatively recently (g. m. pupo, r. lan, and p. r. reeves, proc. natl. acad. sci. usa 97:10567-10572, 2000). there are 33 o-antigen forms in these shigella clones, of which 12 are identical to o antigens of other e. coli strains. we sequenced o-antigen gene clusters from shigella boydii serotypes 4, 5, 6, and 9 and also studied the o53- and o79-antigen gene clusters of e. coli, encoding o antigens identic ... | 2001 | 11598067 |
| rubrerythrin and rubredoxin oxidoreductase in desulfovibrio vulgaris: a novel oxidative stress protection system. | evidence is presented for an alternative to the superoxide dismutase (sod)-catalase oxidative stress defense system in desulfovibrio vulgaris (strain hildenborough). this alternative system consists of the nonheme iron proteins, rubrerythrin (rbr) and rubredoxin oxidoreductase (rbo), the product of the rbo gene (also called desulfoferrodoxin). a deltarbo strain of d. vulgaris was found to be more sensitive to internal superoxide exposure than was the wild type. unlike rbo, expression of plasmid- ... | 2001 | 11114906 |
| characterization and evolution of anthranilate 1,2-dioxygenase from acinetobacter sp. strain adp1. | the two-component anthranilate 1,2-dioxygenase of the bacterium acinetobacter sp. strain adp1 was expressed in escherichia coli and purified to homogeneity. this enzyme converts anthranilate (2-aminobenzoate) to catechol with insertion of both atoms of o(2) and consumption of one nadh. the terminal oxygenase component formed an alpha(3)beta(3) hexamer of 54- and 19-kda subunits. biochemical analyses demonstrated one rieske-type [2fe-2s] center and one mononuclear nonheme iron center in each larg ... | 2001 | 11114907 |
| initiation factor 2 of myxococcus xanthus, a large version of prokaryotic translation initiation factor 2. | we have isolated the structural gene for translation initiation factor if2 (infb) from the myxobacterium myxococcus xanthus. the gene (3.22 kb) encodes a 1,070-residue protein showing extensive homology within its g domain and c terminus to the equivalent regions of if2 from escherichia coli. the protein cross-reacts with antibodies raised against e. coli if2 and was able to complement an e. coli infb mutant. the m. xanthus protein is the largest if2 known to date. this is essentially due to a l ... | 2001 | 11114918 |
| clue to damage recognition by uvrb: residues in the beta-hairpin structure prevent binding to non-damaged dna. | uvrb, the ultimate damage-recognizing component of bacterial nucleotide excision repair, contains a flexible beta-hairpin rich in hydrophobic residues. we describe the properties of uvrb mutants in which these residues have been mutated. the results show that y101 and f108 in the tip of the hairpin are important for the strand-separating activity of uvrb, supporting the model that the beta-hairpin inserts between the two dna strands during the search for dna damage. residues y95 and y96 at the b ... | 2001 | 11689453 |
| structural and mutational studies of the recognition of the arginine trna-specific major identity element, a20, by arginyl-trna synthetase. | arginyl-trna synthetase (argrs) recognizes two major identity elements of trna(arg): a20, located at the outside corner of the l-shaped trna, and c35, the second letter of the anticodon. only a few exceptional organisms, such as the yeast saccharomyces cerevisiae, lack a20 in trna(arg). in the present study, we solved the crystal structure of a typical a20-recognizing argrs from thermus thermophilus at 2.3 a resolution. the structure of the t. thermophilus argrs was found to be similar to that o ... | 2001 | 11698642 |
| brucella abortus genes identified following constitutive growth and macrophage infection. | the chronicity of brucella abortus infection in humans and animals depends on the organism's ability to escape host defenses by gaining entry and surviving inside the macrophage. although no human vaccine exists for brucella, vaccine development in other bacteria has been based on deletions of selective nutritional as well as regulatory systems. our goal is to develop a vaccine for brucella. to further this aim, we have used a green fluorescent protein (gfp) reporter system to identify constitut ... | 2001 | 11705955 |
| ihfa gene of the bacterium myxococcus xanthus and its role in activation of carotenoid genes by blue light. | myxococcus xanthus responds to blue light by producing carotenoids. several regulatory genes are known that participate in the light action mechanism, which leads to the transcriptional activation of the carotenoid genes. we had already reported the isolation of a carotenoid-less, tn5-induced strain (mr508), whose mutant site was unlinked to the indicated regulatory genes. here, we show that omegamr508::tn5 affects all known light-inducible promoters in different ways. it blocks the activation o ... | 2001 | 11133949 |
| crystal structure of thermostable dna photolyase: pyrimidine-dimer recognition mechanism. | dna photolyase is a pyrimidine-dimer repair enzyme that uses visible light. photolyase generally contains two chromophore cofactors. one is a catalytic cofactor directly contributing to the repair of a pyrimidine-dimer. the other is a light-harvesting cofactor, which absorbs visible light and transfers energy to the catalytic cofactor. photolyases are classified according to their second cofactor into either a folate- or deazaflavin-type. the native structures of both types of photolyases have a ... | 2001 | 11707580 |
| gene cluster of rhodothermus marinus high-potential iron-sulfur protein: oxygen oxidoreductase, a caa(3)-type oxidase belonging to the superfamily of heme-copper oxidases. | the respiratory chain of the thermohalophilic bacterium rhodothermus marinus contains an oxygen reductase, which uses hipip (high potential iron-sulfur protein) as an electron donor. the structural genes encoding the four subunits of this hipip:oxygen oxidoreductase were cloned and sequenced. the genes for subunits ii, i, iii, and iv (named rcoxa to rcoxd) are found in this order and seemed to be organized in an operon of at least five genes with a terminator structure a few nucleotides downstre ... | 2001 | 11133964 |
| saturation mutagenesis of 5s rrna in saccharomyces cerevisiae. | rrnas are the central players in the reactions catalyzed by ribosomes, and the individual rrnas are actively involved in different ribosome functions. our previous demonstration that yeast 5s rrna mutants (called mof9) can impact translational reading frame maintenance showed an unexpected function for this ubiquitous biomolecule. at the time, however, the highly repetitive nature of the genes encoding rrnas precluded more detailed genetic and molecular analyses. a new genetic system allows all ... | 2001 | 11713264 |
| overexpression, purification and characterization of recj protein from thermus thermophilus hb8 and its core domain. | a recj homolog was cloned from the extremely thermophilic bacterium thermus themophilus hb8. it encodes a 527 amino acid protein that has 33% identity to escherichia coli recj protein and includes the characteristic motifs conserved among recj homologs. although t.thermophilus recj protein (ttrecj) was expressed as an inclusion body, it was purified in soluble form through denaturation with urea and subsequent refolding steps. limited proteolysis showed that ttrecj has a protease-resistant core ... | 2001 | 11713311 |
| importance of the conserved nucleotides around the trna-like structure of escherichia coli transfer-messenger rna for protein tagging. | a bacterial rna functioning as both trna and mrna, transfer-messenger rna (tmrna) rescues stalled ribosomes and clears the cell of incomplete polypeptides. for function, escherichia coli tmrna requires an elaborate interplay between a trna-like structure and an internal mrna domain that are connected by a 295 nt long compact secondary structure. the trna-like structure is surrounded by 16 unpaired nt, including 10 residues that are >95% conserved among the known 140 tmrna sequences. all these re ... | 2001 | 11713316 |
| behavior of dna fibers stretched by precise meniscus motion control. | a modified dna combing method, which can precisely locate straightened dna fibers on a substrate, has been developed. precise motion control of a dna solution droplet on hydrophobic surfaces has allowed detailed analyses of dna straightening behavior. our method provides a technique for consistently straightening lambda phage dna on a trace of droplet motion, though the straightened dnas had several variations in their alignments. the dependence of the straightened dna frequency upon motion rate ... | 2001 | 11713329 |
| hepatitis c virus 3'x region interacts with human ribosomal proteins. | to identify proteins that can bind the 3' untranslated region (utr) of hepatitis c virus (hcv) we screened human cdna libraries using the saccharomyces cerevisiae three-hybrid system. screening with an rna sequence derived from the 3'-terminal 98 nucleotides (3'x region) of an infectious clone of hcv (h77c) yielded clones of human ribosomal proteins l22, l3, s3, and ml3, a mitochondrial homologue of l3. we performed preliminary characterization of the binding between the 3'x region and these pro ... | 2001 | 11152508 |
| thermostabilization of proteins by diglycerol phosphate, a new compatible solute from the hyperthermophile archaeoglobus fulgidus. | diglycerol phosphate accumulates under salt stress in the archaeon archaeoglobus fulgidus (l. o. martins, r. huber, h. huber, k. o. stetter, m. s. da costa, and h. santos, appl. environ. microbiol. 63:896-902, 1997). this solute was purified after extraction from the cell biomass. in addition, the optically active and the optically inactive (racemic) forms of the compound were synthesized, and the ability of the solute to act as a protecting agent against heating was tested on several proteins d ... | 2000 | 10788369 |
| physical and functional interaction between the eukaryotic orthologs of prokaryotic translation initiation factors if1 and if2. | to initiate protein synthesis, a ribosome with bound initiator methionyl-trna must be assembled at the start codon of an mrna. this process requires the coordinated activities of three translation initiation factors (if) in prokaryotes and at least 12 translation initiation factors in eukaryotes (eif). the factors eif1a and eif5b from eukaryotes show extensive amino acid sequence similarity to the factors if1 and if2 from prokaryotes. by a combination of two-hybrid, coimmunoprecipitation, and in ... | 2000 | 10982835 |
| 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 |
| multiple regulatory mechanisms act on the 5' untranslated region of the s-layer gene from thermus thermophilus hb8. | the role of the 5' untranslated region (5'utr) of the s-layer gene from thermus thermophilus was analyzed through the isolation of delta 5'utr mutants. in these mutants the half-life of spla mrna was strongly reduced and slpa transcription was no longer subjected to growth phase-dependent repression. overproduction and detachment of the external envelopes of the mutants were observed in stationary phase. | 2001 | 11157968 |
| new host-vector system for thermus spp. based on the malate dehydrogenase gene. | a thermus thermophilus hb27 strain was constructed in which the malate dehydrogenase (mdh) gene was deleted. the deltamdh colonies are recognized by a small-colony phenotype. wild-type phenotype is restored by transformation with thermus plasmids or integration vector containing an intact mdh gene. the wild-type phenotype provides a positive selection tool for the introduction of plasmid dna into thermus spp., and because mdh levels can be readily quantified, this host-vector system is a conveni ... | 2001 | 11160114 |
| crystal structure of the lrp-like transcriptional regulator from the archaeon pyrococcus furiosus. | the lrpa protein from the hyperthermophilic archaeon pyrococcus furiosus belongs to the lrp/asnc family of transcriptional regulatory proteins, of which the escherichia coli leucine-responsive regulatory protein is the archetype. its crystal structure has been determined at 2.9 a resolution and is the first for a member of the lrp/asnc family, as well as one of the first for a transcriptional regulator from a hyperthermophile. the structure consists of an n-terminal domain containing a helix-tur ... | 2001 | 11230123 |
| hydrogen peroxide-forming nadh oxidase belonging to the peroxiredoxin oxidoreductase family: existence and physiological role in bacteria. | amphibacillus xylanus and sporolactobacillus inulinus nadh oxidases belonging to the peroxiredoxin oxidoreductase family show extremely high peroxide reductase activity for hydrogen peroxide and alkyl hydroperoxides in the presence of the small disulfide redox protein, ahpc (peroxiredoxin). in order to investigate the distribution of this enzyme system in bacteria, 15 bacterial strains were selected from typical aerobic, facultatively anaerobic, and anaerobic bacteria. ahpc-linked alkyl hydroper ... | 2001 | 11274101 |
| crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and if3. | the small ribosomal subunit is responsible for the decoding of genetic information and plays a key role in the initiation of protein synthesis. we analyzed by x-ray crystallography the structures of three different complexes of the small ribosomal subunit of thermus thermophilus with the a-site inhibitor tetracycline, the universal initiation inhibitor edeine and the c-terminal domain of the translation initiation factor if3. the crystal structure analysis of the complex with tetracycline reveal ... | 2001 | 11296217 |
| conservation of the binding site for the arginine repressor in all bacterial lineages. | the arginine repressor argr/ahrc is a transcription factor universally conserved in bacterial genomes. its recognition signal (the arg box), a weak palindrome, is also conserved between genomes, despite a very low degree of similarity between individual sites within a genome. thus, the arginine repressor is different from two other universal transcription factors - hrca, whose recognition signal is very strongly conserved both within and between genomes, and lexa/dinr, whose signal is strongly c ... | 2001 | 11305941 |
| attached and unattached bacterial communities in a 120-meter corehole in an acidic, crystalline rock aquifer. | the bacteria colonizing geologic core sections (attached) were contrasted with those found suspended in the groundwater (unattached) by examining the microbiology of 16 depth-paired core and groundwater samples using a suite of culture-independent and culture-dependent analyses. one hundred twenty-two meters was continuously cored from a buried chalcopyrite ore hosted in a biotite-quartz-monzonite porphyry at the mineral park mine near kingman, ariz. every fourth 1.5-m core was acquired using mi ... | 2001 | 11319087 |
| oxaloacetate synthesis in the methanarchaeon methanosarcina barkeri: pyruvate carboxylase genes and a putative escherichia coli-type bifunctional biotin protein ligase gene (bpl/bira) exhibit a unique organization. | evidence is presented that, in methanosarcina barkeri oxaloacetate synthesis, an essential and major co(2) fixation reaction is catalyzed by an apparent alpha(4)beta(4)-type acetyl coenzyme a-independent pyruvate carboxylase (pyc), composed of 64.2-kda biotinylated and 52.9-kda atp-binding subunits. the purified enzyme was most active at 70 degrees c, insensitive to aspartate and glutamate, mildly inhibited by alpha-ketoglutarate, and severely inhibited by atp, adp, and excess mg(2+). it showed ... | 2001 | 11371547 |
| spontaneous erythromycin resistance mutation in a 23s rrna gene, rrla, of the extreme thermophile thermus thermophilus ib-21. | spontaneous, erythromycin-resistant mutants of thermus thermophilus ib-21 were isolated and found to carry the mutation a2058g in one of two 23s rrna operons. the heterozygosity of these mutants indicates that a2058g confers a dominant or codominant phenotype in this organism. this mutation provides a valuable tool for the genetic manipulation of the 23s rrna genes of thermus. | 2001 | 11418580 |
| rad54 protein stimulates the postsynaptic phase of rad51 protein-mediated dna strand exchange. | rad54 and rad51 are important proteins for the repair of double-stranded dna breaks by homologous recombination in eukaryotes. as previously shown, rad51 protein forms nucleoprotein filaments on single-stranded dna, and rad54 protein directly interacts with such filaments to enhance synapsis, the homologous pairing with a double-stranded dna partner. here we demonstrate that saccharomyces cerevisiae rad54 protein has an additional role in the postsynaptic phase of dna strand exchange by stimulat ... | 2001 | 11459988 |