| biochemical properties of a high fidelity dna ligase from thermus species ak16d. | nad+-dependent dna ligases from thermophilic bacteria thermus species are highly homologous with amino acid sequence identities ranging from 85 to 98%. thermus species ak16d ligase, the most divergent of the seven thermus isolates collected worldwide, was cloned, expressed in escherichia coli and purified to homogeneity. this thermus ligase is similar to thermus thermophilus hb8 ligase with respect to ph, salt, nad+, divalent cation profiles and steady-state kinetics.however, the former is more ... | 1999 | 9889274 |
| characterization of is1547, a new member of the is900 family in the mycobacterium tuberculosis complex, and its association with is6110. | unlike classically defined insertion sequence (is) elements, which are delimited by their inverted terminal repeats, some is elements do not have inverted terminal repeats. among this group of atypical is elements, is116, is900, is901, and is1110 have been proposed as members of the is900 family of elements, not only because they do not have inverted terminal repeats but also because they share other features such as homologous transposases and particular insertion sites. in this study, we repor ... | 1999 | 9922269 |
| molecular characterization of the nitrite-reducing system of staphylococcus carnosus. | characterization of a nitrite reductase-negative staphylococcus carnosus tn917 mutant led to the identification of the nir operon, which encodes nirbd, the dissimilatory nadh-dependent nitrite reductase; sira, the putative oxidase and chelatase, and sirb, the uroporphyrinogen iii methylase, both of which are necessary for biosynthesis of the siroheme prosthetic group; and nirr, which revealed no convincing similarity to proteins with known functions. we suggest that nirr is essential for nir pro ... | 1999 | 10049379 |
| dissimilatory reduction of fe(iii) and other electron acceptors by a thermus isolate. | a thermophilic bacterium that can use o2, no3-, fe(iii), and s0 as terminal electron acceptors for growth was isolated from groundwater sampled at a 3.2-km depth in a south african gold mine. this organism, designated sa-01, clustered most closely with members of the genus thermus, as determined by 16s rrna gene (rdna) sequence analysis. the 16s rdna sequence of sa-01 was >98% similar to that of thermus strain nmx2 a.1, which was previously isolated by other investigators from a thermal spring i ... | 1999 | 10049886 |
| surface proteins of gram-positive bacteria and mechanisms of their targeting to the cell wall envelope. | the cell wall envelope of gram-positive bacteria is a macromolecular, exoskeletal organelle that is assembled and turned over at designated sites. the cell wall also functions as a surface organelle that allows gram-positive pathogens to interact with their environment, in particular the tissues of the infected host. all of these functions require that surface proteins and enzymes be properly targeted to the cell wall envelope. two basic mechanisms, cell wall sorting and targeting, have been ide ... | 1999 | 10066836 |
| aspartate kinase-independent lysine synthesis in an extremely thermophilic bacterium, thermus thermophilus: lysine is synthesized via alpha-aminoadipic acid not via diaminopimelic acid. | an aspartate kinase-deficient mutant of thermus thermophilus, ak001, was constructed. the mutant strain did not grow in a minimal medium, suggesting that t. thermophilus contains a single aspartate kinase. growth of the mutant strain was restored by addition of both threonine and methionine, while addition of lysine had no detectable effect on growth. to further elucidate the lysine biosynthetic pathway in t. thermophilus, lysine auxotrophic mutants of t. thermophilus were obtained by chemical m ... | 1999 | 10074061 |
| aspartate 205 in the catalytic domain of naphthalene dioxygenase is essential for activity. | the naphthalene dioxygenase enzyme system carries out the first step in the aerobic degradation of naphthalene by pseudomonas sp. strain ncib 9816-4. the crystal structure of naphthalene dioxygenase (b. kauppi, k. lee, e. carredano, r. e. parales, d. t. gibson, h. eklund, and s. ramaswamy, structure 6:571-586, 1998) indicates that aspartate 205 may provide the most direct route of electron transfer between the rieske [2fe-2s] center of one alpha subunit and mononuclear iron in the adjacent alpha ... | 1999 | 10074076 |
| detection and induction of equine infectious anemia virus-specific cytotoxic t-lymphocyte responses by use of recombinant retroviral vectors. | cytotoxic t lymphocytes (ctl) appear to be critical in resolving or reducing the severity of lentivirus infections. retroviral vectors expressing the gag/pr or su protein of the lentivirus equine infectious anemia virus (eiav) were constructed and used to evaluate eiav-specific ctl responses in horses. three promoters, cytomegalovirus, simian virus sv40, and moloney murine sarcoma virus (momsv) long terminal repeat (ltr), were used, and there was considerable variation in their ability to direct ... | 1999 | 10074123 |
| an engineered class i transfer rna with a class ii tertiary fold. | structure-based engineering of the tertiary fold of escherichia coli trna(gln)2 has enabled conversion of this transfer rna to a class ii structure while retaining recognition properties of a class i glutamine trna. the new trna possesses the 20-nt variable stem-loop of thermus thermophilus trna(ser). enlargement of the d-loop appears essential to maintaining a stable tertiary structure in this species, while rearrangement of a base triple in the augmented d-stem is critical for efficient glutam ... | 1999 | 10094311 |
| isolation of rnase h genes that are essential for growth of bacillus subtilis 168. | two genes encoding functional rnase h (ec 3.1.26.4) were isolated from a gram-positive bacterium, bacillus subtilis 168. two dna clones exhibiting rnase h activities both in vivo and in vitro were obtained from a b. subtilis dna library. one (28.2 kda) revealed high similarity to escherichia coli rnase hii, encoded by the rnhb gene. the other (33.9 kda) was designated rnhc and encodes b. subtilis rnase hiii. the b. subtilis genome has an rnha homologue, the product of which has not yet shown rna ... | 1999 | 10094689 |
| nadh-quinone oxidoreductase: psst subunit couples electron transfer from iron-sulfur cluster n2 to quinone. | the proton-translocating nadh-quinone oxidoreductase (ec 1.6.99.3) is the largest and least understood enzyme complex of the respiratory chain. the mammalian mitochondrial enzyme (also called complex i) contains more than 40 subunits, whereas its structurally simpler bacterial counterpart (ndh-1) in paracoccus denitrificans and thermus thermophilus hb-8 consists of 14 subunits. a major unsolved question is the location and mechanism of the terminal electron transfer step from iron-sulfur cluster ... | 1999 | 10097178 |
| distinct affinity of binding sites for s-layer homologous domains in clostridium thermocellum and bacillus anthracis cell envelopes. | binding parameters were determined for the slh (s-layer homologous) domains from the clostridium thermocellum outer layer protein olpb, from the c. thermocellum s-layer protein slpa, and from the bacillus anthracis s-layer proteins ea1 and sap, using cell walls from c. thermocellum and b. anthracis. each slh domain bound to c. thermocellum and b. anthracis cell walls with a different kd, ranging between 7.1 x 10(-7) and 1.8 x 10(-8) m. cell wall binding sites for slh domains displayed different ... | 1999 | 10198008 |
| cis-acting elements responsible for low-temperature-inducible expression of the gene coding for the thermolabile isocitrate dehydrogenase isozyme of a psychrophilic bacterium, vibrio sp. strain abe-1. | transcriptional control of the low-temperature-inducible icdii gene, encoding the thermolabile isocitrate dehydrogenase of a psychrophilic bacterium, vibrio sp. strain abe-1, was found to be mediated in part by a transcriptional silencer locating at nucleotide positions -560 to -526 upstream from the transcription start site of icdii. deletion of the silencer resulted in a 20-fold-increased level of expression of the gene at low temperature (15 degrees c) but not at high temperature (37 degrees ... | 1999 | 10198027 |
| location of translational initiation factor if3 on the small ribosomal subunit. | the location of translational initiation factor if3 bound to the 30s subunit of the thermus thermophilus ribosome has been determined by cryoelectron microscopy. both the 30s.if3 complex and control 30s subunit structures were determined to 27-a resolution. the difference map calculated from the two reconstructions reveals three prominent lobes of positive density. the previously solved crystal structure of if3 fits very well into two of these lobes, whereas the third lobe probably arises from c ... | 1999 | 10200257 |
| purification and characterization of thermotoga maritima endonuclease iv, a thermostable apurinic/apyrimidinic endonuclease and 3'-repair diesterase. | an endonuclease iv homolog was identified as the product of a conceptual open reading frame in the genome of the hyperthermophilic bacterium thermotoga maritima. the t. maritima endonuclease iv gene encodes a 287-amino-acid protein with 32% sequence identity to escherichia coli endonuclease iv. the gene was cloned, and the expressed protein was purified and shown to have enzymatic activities that are characteristic of the endonuclease iv family of dna repair enzymes, including apurinic/apyrimidi ... | 1999 | 10217775 |
| detection of viable listeria monocytogenes with a 5' nuclease pcr assay. | a 5' nuclease assay has been developed to detect viable listeria monocytogenes. the assay targets the hemolysin a (hlya) transcript, which is found only in l. monocytogenes. the single-tube, reverse transcriptase (rt), fluorogenic probe-based assay was formatted by using tth dna polymerase whose activity was modulated by using the manganese-chelating morpholinepropanesulfonic acid (mops) buffer. this assay was quantitative over a 3-log-unit range of template concentrations when tested with an in ... | 1999 | 10224010 |
| sodium-dependent glutamate uptake by an alkaliphilic, thermophilic bacillus strain, ta2.a1. | a strain of bacillus designated ta2.a1, isolated from a thermal spring in te aroha, new zealand, grew optimally at ph 9.2 and 70 degrees c. bacillus strain ta2.a1 utilized glutamate as a sole carbon and energy source for growth, and sodium chloride (>5 mm) was an obligate requirement for growth. growth on glutamate was inhibited by monensin and amiloride, both inhibitors that collapse the sodium gradient (deltapna) across the cell membrane. n, n-dicyclohexylcarbodiimide inhibited the growth of b ... | 1999 | 10322019 |
| three surface layer homology domains at the n terminus of the clostridium cellulovorans major cellulosomal subunit enge. | the gene enge, coding for endoglucanase e, one of the three major subunits of the clostridium cellulovorans cellulosome, has been isolated and sequenced. enge is comprised of an open reading frame (orf) of 3,090 bp and encodes a protein of 1,030 amino acids with a molecular weight of 111,796. the amino acid sequence derived from enge revealed a structure consisting of catalytic and noncatalytic domains. the n-terminal-half region of enge consisted of a signal peptide of 31 amino acid residues an ... | 1999 | 10322032 |
| the type ii pullulanase of thermococcus hydrothermalis: molecular characterization of the gene and expression of the catalytic domain. | the gene encoding a hyperthermostable type ii pullulanase produced by thermococcus hydrothermalis (th-apu) has been isolated. analysis of a total of 5.2 kb of genomic dna has revealed the presence of three open reading frames, one of which (apua) encodes the pullulanase. this enzyme is composed of 1,339 amino acid residues and exhibits a multidomain structure. in addition to a typical n-terminal signal peptide, th-apu possesses a catalytic domain, a domain bearing s-layer homology-like motifs, a ... | 1999 | 10322035 |
| the accessory subunit of xenopus laevis mitochondrial dna polymerase gamma increases processivity of the catalytic subunit of human dna polymerase gamma and is related to class ii aminoacyl-trna synthetases. | peptide sequences obtained from the accessory subunit of xenopus laevis mitochondrial dna (mtdna) polymerase gamma (pol gamma) were used to clone the cdna encoding this protein. amino-terminal sequencing of the mitochondrial protein indicated the presence of a 44-amino-acid mitochondrial targeting sequence, leaving a predicted mature protein with 419 amino acids and a molecular mass of 47.3 kda. this protein is associated with the larger, catalytic subunit in preparations of active mtdna polymer ... | 1999 | 10330144 |
| mannan-degrading enzymes from cellulomonas fimi. | the genes man26a and man2a from cellulomonas fimi encode mannanase 26a (man26a) and beta-mannosidase 2a (man2a), respectively. mature man26a is a secreted, modular protein of 951 amino acids, comprising a catalytic module in family 26 of glycosyl hydrolases, an s-layer homology module, and two modules of unknown function. exposure of man26a produced by escherichia coli to c. fimi protease generates active fragments of the enzyme that correspond to polypeptides with mannanase activity produced by ... | 1999 | 10347049 |
| maltose metabolism in the hyperthermophilic archaeon thermococcus litoralis: purification and characterization of key enzymes. | maltose metabolism was investigated in the hyperthermophilic archaeon thermococcus litoralis. maltose was degraded by the concerted action of 4-alpha-glucanotransferase and maltodextrin phosphorylase (malp). the first enzyme produced glucose and a series of maltodextrins that could be acted upon by malp when the chain length of glucose residues was equal or higher than four, to produce glucose-1-phosphate. phosphoglucomutase activity was also detected in t. litoralis cell extracts. glucose deriv ... | 1999 | 10348846 |
| cd(ii)-responsive and constitutive mutants implicate a novel domain in merr. | expression of the tn21 mercury resistance (mer) operon is controlled by a metal-sensing repressor-activator, merr. when present, merr always binds to the same position on the dna (the operator mero), repressing transcription of the structural genes mertpcad in the absence of hg(ii) and inducing their transcription in the presence of hg(ii). although it has two potential binding sites, the purified merr homodimer binds only one hg(ii) ion, employing cys82 from one monomer and cys117 and cys126 fr ... | 1999 | 10348859 |
| cloning of the glutamyl-trna synthetase (gltx) gene from pseudomonas aeruginosa. | the glutamyl-trna synthetase (gltx) gene from pseudomonas aeruginosa was identified. a plasmid containing a 2.3-kb insert complemented the temperature-sensitive gltx mutation of escherichia coli jp1449, and gltx activity was demonstrated. the inferred amino acid sequence of this gene showed 50.6% identity with gltx from rhizobium meliloti. | 1999 | 10348873 |
| translation elongation factor 1-alpha interacts specifically with the human immunodeficiency virus type 1 gag polyprotein. | human immunodeficiency virus type 1 (hiv-1) gag-encoded proteins play key functions at almost all stages of the viral life cycle. since these functions may require association with cellular factors, the hiv-1 matrix protein (ma) was used as bait in a yeast two-hybrid screen to identify ma-interacting proteins. ma was found to interact with elongation factor 1-alpha (ef1alpha), an essential component of the translation machinery that delivers aminoacyl-trna to ribosomes. ef1alpha was then shown t ... | 1999 | 10364286 |
| nitrate and nitrite control of respiratory nitrate reduction in denitrifying pseudomonas stutzeri by a two-component regulatory system homologous to narxl of escherichia coli. | bacterial denitrification is expressed in response to the concurrent exogenous signals of low-oxygen tension and nitrate or one of its reduction products. the mechanism by which nitrate-dependent gene activation is effected was investigated in the denitrifying bacterium pseudomonas stutzeri atcc 14405. we have identified and isolated from this organism the chromosomal region encoding the two-component sensor-regulator pair narxl and found that it is linked with the narg operon for respiratory ni ... | 1999 | 10368138 |
| the morphological transition of helicobacter pylori cells from spiral to coccoid is preceded by a substantial modification of the cell wall. | the peptidoglycan (murein) of helicobacter pylori has been investigated by high-performance liquid chromatography and mass spectrometric techniques. murein from h. pylori corresponded to the a1gamma chemotype, but the muropeptide elution patterns were substantially different from the one for escherichia coli in that the former produced high proportions of muropeptides with a pentapeptide side chain (about 60 mol%), with gly residues as the c-terminal amino acid (5 to 10 mol%), and with (1-->6)an ... | 1999 | 10368145 |
| heat-inactivated proteins are rescued by the dnak.j-grpe set and clpb chaperones. | functional chaperone cooperation between hsp70 (dnak) and hsp104 (clpb) was demonstrated in vitro. in a eubacterium thermus thermophilus, dnak and dnaj exist as a stable trigonal ring complex (tdnak.j complex) and the dnak gene cluster contains a clpb gene. when substrate proteins were heated at high temperature, none of the chaperones protected them from heat inactivation, but the tdnak.j complex could suppress the aggregation of proteins in an atp- and tgrpe-dependent manner. subsequent incuba ... | 1999 | 10377389 |
| analysis of peptidoglycan structure from vegetative cells of bacillus subtilis 168 and role of pbp 5 in peptidoglycan maturation. | the composition and fine structure of the vegetative cell wall peptidoglycan from bacillus subtilis were determined by analysis of its constituent muropeptides. the structures of 39 muropeptides, representing 97% of the total peptidoglycan, were elucidated. about 99% analyzed muropeptides in b. subtilis vegetative cell peptidoglycan have the free carboxylic group of diaminopimelic acid amidated. anhydromuropeptides and products missing a glucosamine at the nonreducing terminus account for 0.4 an ... | 1999 | 10383963 |
| sulfolobus acidocaldarius inorganic pyrophosphatase: structure, thermostability, and effect of metal ion in an archael pyrophosphatase. | the first crystal structure of an inorganic pyrophosphatase (s-ppase) from an archaebacterium, the thermophile sulfolobus acidocaldarius, has been solved by molecular replacement and refined to an r-factor of 19.7% at 2.7 a. s-ppase is a d3 homohexameric protein with one mg2+ per active site in a position similar to, but not identical with, the first activating metal in mesophilic pyrophosphatases (ppase). in mesophilic ppases, asp65, asp70, and asp102 coordinate the mg2+, while only asp65 and a ... | 1999 | 10386872 |
| the unique chaperone operon of thermotoga maritima: cloning and initial characterization of a functional hsp70 and small heat shock protein. | the hyperthermophilic eubacterium thermotoga maritima possesses an operon encoding an hsp70 molecular chaperone protein and a protein with meaningful homology to the small heat shock protein family of chaperones. this represents the first demonstrated co-operon organization for these two important classes of molecular chaperones. we have cloned and initially characterized these proteins as functional chaperones in vitro: the hsp70 is capable of atp hydrolysis and substrate binding, and the small ... | 1999 | 10400580 |
| characterization of an atypical superoxide dismutase from sinorhizobium meliloti. | sinorhizobium meliloti rm5000 is an aerobic bacterium that can live free in the soil or in symbiosis with the roots of leguminous plants. a single detectable superoxide dismutase (sod) was found in free-living growth conditions. the corresponding gene was isolated from a genomic library by using a sod fragment amplified by pcr from degenerate primers as a probe. the soda gene was located in the chromosome. it is transcribed monocistronically and encodes a 200-amino-acid protein with a theoretica ... | 1999 | 10419947 |
| in thermoanaerobacterium thermosulfurigenes em1 s-layer homology domains do not attach to peptidoglycan. | three exocellular enzymes of thermoanaerobacterium thermosulfurigenes em1 possess a c-terminal triplicated sequence related to a domain of bacterial cell surface proteins (s-layer proteins). at least one copy of this sequence, named the slh (for s-layer homology) domain, is also present at the n terminus of the s-layer protein of this bacterium. the hypothesis that slh domains serve to anchor proteins to the cell surface was investigated by using the slh domain-containing xylanase. this enzyme w ... | 1999 | 10438774 |
| an inducible human immunodeficiency virus type 1 (hiv-1) vector which effectively suppresses hiv-1 replication. | recently, gene therapy vectors based upon the human immunodeficiency virus type 1 (hiv-1) genome have been developed. here, we create an hiv-1 vector which is defective for all hiv-1 genes, but which maintains cis-acting elements required for efficient packaging, infection, and expression. in t cells transduced by this vector, vector expression is low but efficiently induced following hiv-1 infection. remarkably, although the hiv-1 vector does not contain specific anti-hiv-1 therapeutic genes, t ... | 1999 | 10438857 |
| the accessory subunit of mtdna polymerase shares structural homology with aminoacyl-trna synthetases: implications for a dual role as a primer recognition factor and processivity clamp. | the accessory subunit of the heterodimeric mtdna polymerase (polgamma) from drosophila embryos is required to maintain the structural integrity or catalytic efficiency of the holoenzyme. cdnas for the accessory subunit from drosophila, man, mouse, and rat have been identified, and comparative sequence alignment reveals that the c-terminal region of about 120 aa is the most conserved. furthermore, we demonstrate that the accessory subunit of animal polgamma has both sequence and structural simila ... | 1999 | 10449726 |
| active-site mutations in the xrn1p exoribonuclease of saccharomyces cerevisiae reveal a specific role in meiosis. | xrn1p of saccharomyces cerevisiae is a major cytoplasmic rna turnover exonuclease which is evolutionarily conserved from yeasts to mammals. deletion of the xrn1 gene causes pleiotropic phenotypes, which have been interpreted as indirect consequences of the rna turnover defect. by sequence comparisons, we have identified three loosely defined, common 5'-3' exonuclease motifs. the significance of motif ii has been confirmed by mutant analysis with xrn1p. the amino acid changes d206a and d208a abol ... | 1999 | 10454540 |
| comparison of the d-glutamate-adding enzymes from selected gram-positive and gram-negative bacteria. | the biochemical properties of the d-glutamate-adding enzymes (murd) from escherichia coli, haemophilus influenzae, enterococcus faecalis, and staphylococcus aureus were investigated to detect any differences in the activity of this enzyme between gram-positive and gram-negative bacteria. the genes (murd) that encode these enzymes were cloned into pmal-c2 fusion vector and overexpressed as maltose-binding protein-murd fusion proteins. each fusion protein was purified to homogeneity by affinity to ... | 1999 | 10464212 |
| marine bacterial isolates display diverse responses to uv-b radiation. | the molecular and biological consequences of uv-b radiation were investigated by studying five species of marine bacteria and one enteric bacterium. laboratory cultures were exposed to an artificial uv-b source and subjected to various post-uv irradiation treatments. significant differences in survival subsequent to uv-b radiation were observed among the isolates, as measured by culturable counts. uv-b-induced dna photodamage was investigated by using a highly specific radioimmunoassay to measur ... | 1999 | 10473381 |
| cloning of the gene encoding a novel thermostable alpha-galactosidase from thermus brockianus iti360. | an alpha-galactosidase gene from thermus brockianus iti360 was cloned, sequenced, and expressed in escherichia coli, and the recombinant protein was purified. the gene, designated agat, codes for a 476-residue polypeptide with a calculated molecular mass of 53, 810 da. the native structure of the recombinant enzyme (agat) was estimated to be a tetramer. agat displays amino acid sequence similarity to the alpha-galactosidases of thermotoga neapolitana and thermotoga maritima and a low-level seque ... | 1999 | 10473401 |
| p1 parb domain structure includes two independent multimerization domains. | parb is one of two p1-encoded proteins that are required for active partition of the p1 prophage in escherichia coli. to probe the native domain structure of parb, we performed limited proteolytic digestions of full-length parb, as well as of several n-terminal and c-terminal deletion fragments of parb. the c-terminal 140 amino acids of parb form a very trypsin-resistant domain. in contrast, the n terminus is more susceptible to proteolysis, suggesting that it forms a less stably folded domain o ... | 1999 | 10498700 |
| crystal structure of the dna nucleotide excision repair enzyme uvrb from thermus thermophilus. | nucleotide excision repair (ner) is the most important dna-repair mechanism in living organisms. in prokaryotes, three enzymes forming the uvrabc system initiate ner of a variety of structurally different dna lesions. uvrb, the central component of this system, is responsible for the ultimate dna damage recognition and participates in the incision of the damaged dna strand. the crystal structure of thermus thermophilus uvrb reveals a core that is structurally similar to core regions found in hel ... | 1999 | 10518516 |
| detection of human herpesvirus 6 by reverse transcription-pcr. | the role of human herpesvirus 6 (hhv-6) in disease beyond primary infection remains unclear. we have developed and validated a new reverse transcription-pcr (rt-pcr) assay for hhv-6 that can determine the presence of hhv-6 in clinical specimens and differentiate between latent and replicating virus. peripheral blood mononuclear cells from 109 children were evaluated for hhv-6 by rt-pcr, dna pcr, and viral culture. of these samples, 106 were suitable for analysis. a total of 20 samples were posit ... | 1999 | 10523572 |
| loss of cytochrome c oxidase activity and acquisition of resistance to quinone analogs in a laccase-positive variant of azospirillum lipoferum. | laccase, a p-diphenol oxidase typical of plants and fungi, has been found recently in a proteobacterium, azospirillum lipoferum. laccase activity was detected in both a natural isolate and an in vitro-obtained phase variant that originated from the laccase-negative wild type. in this study, the electron transport systems of the laccase-positive variant and its parental laccase-negative forms were compared. during exponential (but not stationary) growth under fully aerobic (but not under microaer ... | 1999 | 10542175 |
| specific inhibition of the eubacterial dna ligase by arylamino compounds. | all known dna ligases catalyze the formation of a phosphodiester linkage between adjacent termini in double-stranded dna via very similar mechanisms. the ligase family can, however, be divided into two classes: eubacterial ligases, which require nad(+) as a cofactor, and other ligases, from viruses, archaea, and eukaryotes, which use atp. drugs that discriminate between dna ligases from different sources may have antieubacterial activity. we now report that a group of arylamino compounds, includ ... | 1999 | 10543760 |
| improving proteolytic cleavage at the 3a/3b site of the hepatitis a virus polyprotein impairs processing and particle formation, and the impairment can be complemented in trans by 3ab and 3abc. | the orchestrated liberation of viral proteins by 3c(pro)-mediated proteolysis is pivotal for gene expression by picornaviruses. proteolytic processing is regulated either by the amino acid sequence at the cleavage site of the substrate or by cofactors covalently or noncovalently linked to the viral proteinase. to determine the role of the amino acid sequence at cleavage sites 3a/3b and 3b/3c that are essential for the liberation of 3c(pro) from its precursors and to assess the function of the st ... | 1999 | 10559299 |
| determination of bovine rotavirus g and p serotypes in italy by pcr. | determination of the g and p serotypes of group a bovine rotaviruses from 149 samples of feces or intestinal contents collected from calves showing clinical signs of neonatal diarrhea was performed by a nested reverse transcription-pcr typing assay. the g6 serotype was the most prevalent, accounting for viruses in 55.7% of the samples; viruses of the g10 and g8 serotypes were found in 34.9 and 4.7% of the samples, respectively. the virus in one sample (0.7%) was not classified due to concomitant ... | 1999 | 10565900 |
| genome sequence of the radioresistant bacterium deinococcus radiodurans r1. | the complete genome sequence of the radiation-resistant bacterium deinococcus radiodurans r1 is composed of two chromosomes (2,648,638 and 412,348 base pairs), a megaplasmid (177,466 base pairs), and a small plasmid (45,704 base pairs), yielding a total genome of 3,284, 156 base pairs. multiple components distributed on the chromosomes and megaplasmid that contribute to the ability of d. radiodurans to survive under conditions of starvation, oxidative stress, and high amounts of dna damage were ... | 1999 | 10567266 |
| structural distribution of stability in a thermophilic enzyme. | stability parameters for individual residues in thermus thermophilus cysteine-free rnase h were determined by native state hydrogen exchange, thus providing a unique comparison of regional thermodynamics between thermophilic and mesophilic homologues. the general distribution of stability in the thermophilic protein is similar to that of its mesophilic homologue, with a proportional increase in stability for almost all residues. as a consequence, the residue-specific stabilities of the two prote ... | 1999 | 10570131 |
| sequential mechanism of solubilization and refolding of stable protein aggregates by a bichaperone network. | a major activity of molecular chaperones is to prevent aggregation and refold misfolded proteins. however, when allowed to form, protein aggregates are refolded poorly by most chaperones. we show here that the sequential action of two escherichia coli chaperone systems, clpb and dnak-dnaj-grpe, can efficiently solubilize excess amounts of protein aggregates and refold them into active proteins. measurements of aggregate turbidity, congo red, and 4,4'-dianilino-1, 1'-binaphthyl-5,5'-disulfonic ac ... | 1999 | 10570141 |
| ligase-based detection of mononucleotide repeat sequences. | up to 15% of all colorectal cancers are considered to be replication error positive (rer(+)) and contain mutations at hundreds of thousands of microsatellite repeat sequences. recently, a number of intragenic mononucleotide repeat sequences have been demonstrated to be targets for inactivating genes in rer(+)colorectal tumors. in this study, thermostable dna ligases were tested for the ability to detect alterations in microsatellite sequences in colon tumor samples. ligation profiles on mononucl ... | 1999 | 10572192 |
| cloning, overexpression, and mutagenesis of the sporobolomyces salmonicolor aku4429 gene encoding a new aldehyde reductase, which catalyzes the stereoselective reduction of ethyl 4-chloro-3-oxobutanoate to ethyl (s)-4-chloro-3-hydroxybutanoate. | we cloned and sequenced the gene encoding an nadph-dependent aldehyde reductase (arii) in sporobolomyces salmonicolor aku4429, which reduces ethyl 4-chloro-3-oxobutanoate (4-cobe) to ethyl (s)-4-chloro-3-hydroxybutanoate. the arii gene is 1,032 bp long, is interrupted by four introns, and encodes a 37,315-da polypeptide. the deduced amino acid sequence exhibited significant levels of similarity to the amino acid sequences of members of the mammalian 3beta-hydroxysteroid dehydrogenase-plant dihyd ... | 1999 | 10583966 |
| stress genes and proteins in the archaea. | the field covered in this review is new; the first sequence of a gene encoding the molecular chaperone hsp70 and the first description of a chaperonin in the archaea were reported in 1991. these findings boosted research in other areas beyond the archaea that were directly relevant to bacteria and eukaryotes, for example, stress gene regulation, the structure-function relationship of the chaperonin complex, protein-based molecular phylogeny of organisms and eukaryotic-cell organelles, molecular ... | 1999 | 10585970 |
| the small ribosomal subunit from thermus thermophilus at 4.5 a resolution: pattern fittings and the identification of a functional site. | the electron density map of the small ribosomal subunit from thermus thermophilus, constructed at 4.5 a resolution, shows the recognizable morphology of this particle, as well as structural features that were interpreted as ribosomal rna and proteins. unbiased assignments, carried out by quantitative covalent binding of heavy atom compounds at predetermined sites, led to the localization of the surface of the ribosomal protein s13 at a position compatible with previous assignments, whereas the s ... | 1999 | 10588692 |
| 5s ribosomal rna database y2k. | this paper presents the updated version (y2k) of the database of ribosomal 5s ribonucleic acids (5s rrna) and their genes (5s rdna), http://rose.man/poznan.pl/5sdata/index.html. this edition of the database contains 1985primary structures of 5s rrna and 5s rdna. they include 60 archaebacterial, 470 eubacterial, 63 plastid, nine mitochondrial and 1383 eukaryotic sequences. the nucleotide sequences of the 5s rrnas or 5s rdnas are divided according to the taxonomic position of the source organisms. | 2000 | 10592212 |
| aminoacyl-trna synthetases database y2k. | the aminoacyl-trna synthetases (aars) are a diverse group of enzymes that ensure the fidelity of transfer of genetic information from dna into protein. they catalyse the attachment of amino acids to transfer rnas and thereby establish the rules of the genetic code by virtue of matching the nucleotide triplet of the anticodon with its cognate amino acid. currently, 818 aars primary structures have been reported from archaebacteria, eubacteria, mitochondria, chloro-plasts and eukaryotic cells. the ... | 2000 | 10592262 |
| structural and functional analyses of the secondary cell wall polymer of bacillus sphaericus ccm 2177 that serves as an s-layer-specific anchor. | sacculi of bacillus sphaericus ccm 2177 contain a secondary cell wall polymer which was completely extracted with 48% hydrofluoric acid. nuclear magnetic resonance analysis showed that the polymer is composed of repeating units, as follows: -->3)-[4, 6-o-(1-carboxyethylidene)]( approximately 0. 5)-beta-d-manpnac-(1-->4)-beta-d-glcpnac-(1-->. the n-terminal part of the s-layer protein carrying s-layer homologous motifs recognizes this polymer as a binding site. | 1999 | 10601228 |
| the heme-copper oxidases of thermus thermophilus catalyze the reduction of nitric oxide: evolutionary implications. | we show that the heme-copper terminal oxidases of thermus thermophilus (called ba(3) and caa(3)) are able to catalyze the reduction of nitric oxide (no) to nitrous oxide (n(2)o) under reducing anaerobic conditions. the rate of no consumption and n(2)o production were found to be linearly dependent on enzyme concentration, and activity was abolished by enzyme denaturation. thus, contrary to the eukaryotic enzyme, both t. thermophilus oxidases display a no reductase activity (3.0 +/- 0.7 mol no/mo ... | 1999 | 10611279 |
| identification and characterization of immunoglobulin g in blood as a major inhibitor of diagnostic pcr. | a major inhibitor of diagnostic pcr in human plasma was identified and the mechanism of inhibition was characterized. human blood was divided by centrifugation into buffy coat, plasma, platelets, and erythrocytes. all these blood fractions were found to be highly inhibitory to a standardized pcr mixture containing the thermostable dna polymerase amplitaq gold. pcr inhibitors in human plasma were purified by chromatographic procedures and were characterized by a process of elimination, so that th ... | 2000 | 10618113 |
| functional analysis of the small component of the 4-hydroxyphenylacetate 3-monooxygenase of escherichia coli w: a prototype of a new flavin:nad(p)h reductase subfamily. | escherichia coli w uses the aromatic compound 4-hydroxyphenylacetate (4-hpa) as a sole source of carbon and energy for growth. the monooxygenase which converts 4-hpa into 3,4-dihydroxyphenylacetate, the first intermediate of the pathway, consists of two components, hpab (58.7 kda) and hpac (18.6 kda), encoded by the hpab and hpac genes, respectively, that form a single transcription unit. overproduction of the small hpac component in e. coli k-12 cells has facilitated the purification of the pro ... | 2000 | 10633095 |
| flexibility, conformational diversity and two dimerization modes in complexes of ribosomal protein l12. | protein l12, the only multicopy component of the ribosome, is presumed to be involved in the binding of translation factors, stimulating factor-dependent gtp hydrolysis. crystal structures of l12 from thermotogamaritima have been solved in two space groups by the multiple anomalous dispersion method and refined at 2.4 and 2.0 a resolution. in both crystal forms, an asymmetric unit comprises two full-length l12 molecules and two n-terminal l12 fragments that are associated in a specific, hetero-t ... | 2000 | 10637222 |
| active site constraints in the hydrolysis reaction catalyzed by bacterial rnase p: analysis of precursor trnas with a single 3'-s-phosphorothiolate internucleotide linkage. | endonucleolytic processing of precursor trnas (ptrnas) by rnase p yields 3'-oh and 5'-phosphate termini, and at least two metal ions are thought to be essential for catalysis. to determine if the hydrolysis reaction catalyzed by bacterial rnase p (rnas) involves stabilization of the 3'-oxyanion leaving group by direct coordination to one of the catalytic metal ions, ptrna substrates with single 3'- s -phosphorothiolate linkages at the rnase p cleavage site were synthesized. with a 3'- s -phospho ... | 2000 | 10637323 |
| optimised ligation of oligonucleotides by thermal ligases: comparison of thermus scotoductus and rhodothermus marinus dna ligases to other thermophilic ligases. | we describe the characterisation of four thermo-stable nad(+)-dependent dna ligases, from thermus thermophilus (tth), thermus scotoductus (ts), rhodothermus marinus (rm) and thermus aquaticus (taq), by an assay which measures ligation rate and mismatch discrimination. complete libraries of octa-, nona- and decanucleotides were used as substrates. the assay comprised the polymerisation of oligo-nucleotides initiated from a 17 base 'primer', using m13mp18 ssdna as template. polymers of ligation pr ... | 2000 | 10637340 |
| structure and expression of the human p68 rna helicase gene. | nuclear dead box protein p68 is immunologically related to sv40 large tumour antigen and both proteins possess rna helicase activity. in this report, we describe the structural organisation of the human p68 gene and aspects of the regulation of its expression. northern blot and primer extension analyses indicate that, although its level is variable, the p68 rna helicase appears to be expressed from a single transcription start site in all tissues tested. sequence analysis revealed that the p68 p ... | 2000 | 10648785 |
| 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 |
| 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 |
| 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 |
| modified constructs of the trna tpsic domain to probe substrate conformational requirements of m(1)a(58) and m(5)u(54) trna methyltransferases. | the tpsic stem and loop (tsl) of trna contains highly conserved nucleoside modifications, m(5)c(49), t(54), psi(55)and m(1)a(58). u(54)is methylated to m(5)u (t) by m(5)u(54)methyltransferase (rumt); a(58)is methylated to m(1)a by m(1)a(58)trna methyltransferase (ramt). rumt recognizes and methylates a minimal tsl heptadecamer and ramt has previously been reported to recognize and methylate the 3'-half of the trna molecule. we report that ramt can recognize and methylate a tsl heptadecamer. to b ... | 2000 | 10684932 |
| ligation reaction specificities of an nad(+)-dependent dna ligase from the hyperthermophile aquifex aeolicus. | an nad(+)-dependent dna ligase from the hyperthermophilic bacterium aquifex aeolicus was cloned, expressed in escherichia coli and purified to homogeneity. the enzyme is most active in slightly alkaline ph conditions with either mg(2+)or mn(2+)as the metal cofactor. ca(2+)and ni(2+)mainly support formation of dna-adenylate intermediates. the catalytic cycle is characterized by a low k (cat)value of 2 min(-1)with concomitant accumulation of the dna - adenylate intermediate when mg(2+)is used as t ... | 2000 | 10684941 |
| evolution of arginine biosynthesis in the bacterial domain: novel gene-enzyme relationships from psychrophilic moritella strains (vibrionaceae) and evolutionary significance of n-alpha-acetyl ornithinase. | in the arginine biosynthetic pathway of the vast majority of prokaryotes, the formation of ornithine is catalyzed by an enzyme transferring the acetyl group of n-alpha-acetylornithine to glutamate (ornithine acetyltransferase [oatase]) (argj encoded). only two exceptions had been reported-the enterobacteriaceae and myxococcus xanthus (members of the gamma and delta groups of the class proteobacteria, respectively)-in which ornithine is produced from n-alpha-acetylornithine by a deacylase, acetyl ... | 2000 | 10692366 |
| 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 |
| metabolic engineering of saccharomyces cerevisiae. | comprehensive knowledge regarding saccharomyces cerevisiae has accumulated over time, and today s. cerevisiae serves as a widley used biotechnological production organism as well as a eukaryotic model system. the high transformation efficiency, in addition to the availability of the complete yeast genome sequence, has facilitated genetic manipulation of this microorganism, and new approaches are constantly being taken to metabolicially engineer this organism in order to suit specific needs. in t ... | 2000 | 10704473 |
| 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 |
| a missense mutation in the nuclear gene coding for the mitochondrial aspartyl-trna synthetase suppresses a mitochondrial trna(asp) mutation. | the nuclear suppressor allele nsm3 in strain ff1210-6c/170-e22 (e22), which suppresses a mutation of the yeast mitochondrial trna(asp)gene in saccharomyces cerevisiae, was cloned and identified. to isolate the nsm3 allele, a genomic dna library using the vector yep13 was constructed from strain e22. nine yep13 recombinant plasmids were isolated and shown to suppress the mutation in the mitochondrial trna(asp)gene. these nine plasmids carry a common 4. 5-kb chromosomal dna fragment which contains ... | 2000 | 10710420 |
| 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 |
| two nitrate/nitrite transporters are encoded within the mobilizable plasmid for nitrate respiration of thermus thermophilus hb8. | thermus thermophilus hb8 can grow anaerobically by using a membrane-bound nitrate reductase to catalyze the reduction of nitrate as a final electron acceptor in respiration. in contrast to other denitrifiers, the nitrite produced does not continue the reduction pathway but accumulates in the growth medium after its active extrusion from the cell. we describe the presence of two genes, nark1 and nark2, downstream of the nitrate reductase-encoding gene cluster (nar) that code for two homologues to ... | 2000 | 10735860 |
| expression of a heterologous glutamate dehydrogenase gene in lactococcus lactis highly improves the conversion of amino acids to aroma compounds. | the first step of amino acid degradation in lactococci is a transamination, which requires an alpha-keto acid as the amino group acceptor. we have previously shown that the level of available alpha-keto acid in semihard cheese is the first limiting factor for conversion of amino acids to aroma compounds, since aroma formation is greatly enhanced by adding alpha-ketoglutarate to cheese curd. in this study we introduced a heterologous catabolic glutamate dehydrogenase (gdh) gene into lactococcus l ... | 2000 | 10742211 |
| 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 |
| genome plasticity among related ++lactococcus strains: identification of genetic events associated with macrorestriction polymorphisms. | the genomic diversity of nine strains of the lactococcus lactis subsp. cremoris (ncdo712, ncdo505, ncdo2031, ncdo763, mms36, c2, lm0230, lm2301, and mg1363) was studied by macrorestriction enzyme analysis using pulsed-field gel electrophoresis. these strains were considered adequate for the investigation of genomic plasticity because they have been described as belonging to the same genetic lineage. comparison of apai and smai genome fingerprints of each strain revealed the presence of several m ... | 2000 | 10762249 |
| 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 |
| 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 |
| 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 |
| 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 |
| conversion of a c type cytochrome to a b type that spontaneously forms in vitro from apo protein and heme: implications for c type cytochrome biogenesis and folding. | cytochrome c(552) from hydrogenobacter thermophilus, a thermophilic bacterium, has been converted into a b type cytochrome, after mutagenesis of both heme-binding cysteines to alanine and expression in the cytoplasm of escherichia coli. the b type variant is less stable, with the guanidine hydrochloride unfolding midpoint occurring at a concentration 2 m lower than for the wild-type protein. the reduction potential is 75 mv lower than that of the recombinant wild-type protein. the heme can be re ... | 2000 | 10792037 |
| the 2 a crystal structure of leucyl-trna synthetase and its complex with a leucyl-adenylate analogue. | leucyl-, isoleucyl- and valyl-trna synthetases are closely related large monomeric class i synthetases. each contains a homologous insertion domain of approximately 200 residues, which is thought to permit them to hydrolyse ('edit') cognate trna that has been mischarged with a chemically similar but non-cognate amino acid. we describe the first crystal structure of a leucyl-trna synthetase, from the hyperthermophile thermus thermophilus, at 2.0 a resolution. the overall architecture is similar t ... | 2000 | 10811626 |
| performance of the cobas amplicor hcv monitor test, version 2.0, an automated reverse transcription-pcr quantitative system for hepatitis c virus load determination. | a clinical evaluation of an automated quantitative pcr assay, the cobas amplicor hcv monitor test, version 2.0 (v2.0), was carried out to assess the performance of this test in comparison with that of the previous, manual version, the amplicor hcv monitor test, and with that of nested pcr. serial dilutions of serum samples infected with genotype 1b, 2a, or 3, as well as synthetic rna transcripts and serum samples derived from 87 patients with chronic hepatitis c and infected with genotype 1a, 1b ... | 2000 | 10834978 |
| three-dimensional cryo-electron microscopy localization of ef2 in the saccharomyces cerevisiae 80s ribosome at 17.5 a resolution. | using a sordarin derivative, an antifungal drug, it was possible to determine the structure of a eukaryotic ribosome small middle dotef2 complex at 17.5 a resolution by three-dimensional (3d) cryo-electron microscopy. ef2 is directly visible in the 3d map and the overall arrangement of the complex from saccharomyces cerevisiae corresponds to that previously seen in escherichia coli. however, pronounced differences were found in two prominent regions. first, in the yeast system the interaction be ... | 2000 | 10835368 |
| the location of protein s8 and surrounding elements of 16s rrna in the 70s ribosome from combined use of directed hydroxyl radical probing and x-ray crystallography. | ribosomal protein s8, which is essential for the assembly of the central domain of 16s rrna, is one of the most thoroughly studied rna-binding proteins. to map its surrounding rna in the ribosome, we carried out directed hydroxyl radical probing of 16s rrna using fe(ii) tethered to nine different positions on the surface of protein s8 in 70s ribosomes. hydroxyl radical-induced cleavage was observed near the classical s8-binding site in the 620 stem, and flanking the other s8-footprinted regions ... | 2000 | 10836793 |
| peptide deformylase in staphylococcus aureus: resistance to inhibition is mediated by mutations in the formyltransferase gene. | peptide deformylase, a bacterial enzyme, represents a novel target for antibiotic discovery. two deformylase homologs, defa and defb, were identified in staphylococcus aureus. the defa homolog, located upstream of the transformylase gene, was identified by genomic analysis and was cloned from chromosomal dna by pcr. a distinct homolog, defb, was cloned from an s. aureus genomic library by complementation of the arabinose-dependent phenotype of a p(bad)-def escherichia coli strain grown under ara ... | 2000 | 10858337 |
| characterization of an atp-dependent dna ligase from the thermophilic archaeon methanobacterium thermoautotrophicum. | we report the production, purification and characterization of a dna ligase encoded by the thermophilic archaeon methanobacterium thermoautotrophicum. the 561 amino acid mth: ligase catalyzed strand-joining on a singly nicked dna in the presence of a divalent cation (magnesium, manganese or cobalt) and atp (k(m) 1.1 microm). datp can substitute for atp, but ctp, gtp, utp and nad(+) cannot. mth: ligase activity is thermophilic in vitro, with optimal nick-joining at 60 degrees c. mutational analys ... | 2000 | 10871342 |
| importance of discriminator base stacking interactions: molecular dynamics analysis of a73 microhelix(ala) variants. | transfer of alanine from escherichia coli alanyl-trna synthetase (alars) to rna minihelices that mimic the amino acid acceptor stem of trna(ala) has been shown, by analysis of variant minihelix aminoacylation activities, to involve a transition state sensitive to changes in the 'discriminator' base at position 73. solution nmr has indicated that this single-stranded nucleotide is predominantly stacked onto g1 of the first base pair of the alanine acceptor stem helix. we report the activity of a ... | 2000 | 10871402 |
| 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 |
| crystal structure of cystalysin from treponema denticola: a pyridoxal 5'-phosphate-dependent protein acting as a haemolytic enzyme. | cystalysin is a c(beta)-s(gamma) lyase from the oral pathogen treponema denticola catabolyzing l-cysteine to produce pyruvate, ammonia and h(2)s. with its ability to induce cell lysis, cystalysin represents a new class of pyridoxal 5'-phosphate (plp)-dependent virulence factors. the crystal structure of cystalysin was solved at 1.9 a resolution and revealed a folding and quaternary arrangement similar to aminotransferases. based on the active site architecture, a detailed catalytic mechanism is ... | 2000 | 10880431 |
| arginines 29 and 59 of elongation factor g are important for gtp hydrolysis or translocation on the ribosome. | gtp hydrolysis by elongation factor g (ef-g) is essential for the translocation step in protein elongation. the low intrinsic gtpase activity of ef-g is strongly stimulated by the ribosome. here we show that a conserved arginine, r29, of escherichia coli ef-g is crucial for gtp hydrolysis on the ribosome, but not for gtp binding or ribosome interaction, suggesting that it may be directly involved in catalysis. another conserved arginine, r59, which is homologous to the catalytic arginine of g(al ... | 2000 | 10880458 |
| 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 carboxy-terminal 16-amino-acid region of sigma(38) of escherichia coli is important for transcription under high-salt conditions and sigma activities in vivo. | sigma(38) (or sigma(s), the rpos gene product) is a sigma subunit of rna polymerase in escherichia coli and directs transcription from a number of stationary-phase promoters as well as osmotically inducible promoters. in this study, we analyzed the function of the carboxy-terminal 16-amino-acid region of sigma(38) (residues 315 to 330), which is well conserved among the rpos gene products of enteric bacterial species. truncation of this region was shown to result in the loss of sigma activity in ... | 2000 | 10913098 |
| sequencing, cloning, and high-level expression of the pfp gene, encoding a pp(i)-dependent phosphofructokinase from the extremely thermophilic eubacterium dictyoglomus thermophilum. | the sequencing, cloning, and expression of the pfp gene from dictyoglomus thermophilum, which consists of 1,041 bp and encodes a pyrophosphate-dependent phosphofructokinase, are described. a phylogenetic analysis indicates that the enzyme is closely related to the pyrophosphate-dependent enzyme from thermoproteus tenax. the recombinant and native enzymes share a high degree of similarity for most properties examined. | 2000 | 10913106 |
| 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 |
| 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 |