| structure determination of two new amino acid-containing derivatives of adenosine from trna of thermophilic bacteria and archaea. | two new nucleosides have been identified in unfractionated transfer rna of two thermophilic bacteria, thermodesulfobacterium commune, and thermotoga maritima, six hyperthermophilic archaea, including pyrobaculum islandicum, pyrococcus furiosus and thermococcus sp. and two mesophilic archaea, methanococcus vannielii and methanolobus tindarius. structures were determined primarily by mass spectrometry, as 3-hydroxy-n-[[(9-beta-d-ribofuranosyl-9h-purin-6- yl)amino]carbonyl]norvaline, (hn6a), struct ... | 1992 | 1280806 |
| x-ray crystal structures of the oxidized and reduced forms of the rubredoxin from the marine hyperthermophilic archaebacterium pyrococcus furiosus. | the structures of the oxidized and reduced forms of the rubredoxin from the archaebacterium, pyrococcus furiosus, an organism that grows optimally at 100 degrees c, have been determined by x-ray crystallography to a resolution of 1.8 a. crystals of this rubredoxin grow in space group p2(1)2(1)2(1) with room temperature cell dimensions a = 34.6 a, b = 35.5 a, and c = 44.4 a. initial phases were determined by the method of molecular replacement using the oxidized form of the rubredoxin from the me ... | 1992 | 1303768 |
| solution-state structure by nmr of zinc-substituted rubredoxin from the marine hyperthermophilic archaebacterium pyrococcus furiosus. | the three-dimensional solution-state structure is reported for the zinc-substituted form of rubredoxin (rd) from the marine hyperthermophilic archaebacterium pyrococcus furiosus, an organism that grows optimally at 100 degrees c. structures were generated with dspace by a hybrid distance geometry (dg)-based simulated annealing (sa) approach that employed 403 nuclear overhauser effect (noe)-derived interproton distance restraints, including 67 interresidue, 124 sequential (i-j = 1), 75 medium-ran ... | 1992 | 1303769 |
| comparison of the x-ray structure of native rubredoxin from pyrococcus furiosus with the nmr structure of the zinc-substituted protein. | the three-dimensional x-ray structures of the oxidized and reduced forms of rubredoxin from pyrococcus furiosus, determined at -161 degrees c, and the nmr structure of the zinc-substituted protein, determined in solution at 45 degrees c, are compared. the nmr and x-ray structures, which were determined independently, are very similar and lead to similar conclusions regarding the interactions that confer hyperthermostability. | 1992 | 1303770 |
| hydrogen-oxidizing electron transport components in the hyperthermophilic archaebacterium pyrodictium brockii. | the hyperthermophilic archaebacterium pyrodictium brockii grows optimally at 105 degrees c by a form of metabolism known as hydrogen-sulfur autotrophy, which is characterized by the oxidation of h2 by s0 to produce atp and h2s. uv-irradiated membranes were not able to carry out the hydrogen-dependent reduction of sulfur. however, the activity could be restored by the addition of ubiquinone q10 or ubiquinone q6 to the uv-damaged membranes. a quinone with thin-layer chromatography migration proper ... | 1992 | 1309514 |
| the glna gene of the extremely thermophilic eubacterium thermotoga maritima: cloning, primary structure, and expression in escherichia coli. | the structural gene (glna) encoding the glutamine synthetase (gs) of the extremely thermophilic eubacterium thermotoga maritima has been cloned on a 6.0 kb hindiii dna fragment. sequencing of the region containing the glna gene (1444 bp) showed an orf encoding a polypeptide (439 residues) with an estimated mass of 50,088 da, which shared significant homology with the gsi sequences of other bacteria (escherichia coli, bacillus subtilis) and archaea (pyrococcus woesei, sulfolobus solfataricus). th ... | 1992 | 1348781 |
| quantitative measurement of small through-hydrogen-bond and 'through-space' 1h-113cd and 1h-199hg j couplings in metal-substituted rubredoxin from pyrococcus furiosus. | a method is described for measurement of small unresolvable heteronuclear j couplings. the method is based on quantitative analysis of a phase-purged heteronuclear spin-echo difference spectrum, and is demonstrated for measuring 1h-113cd and 1h-199hg j couplings in metal-substituted rubredoxin (m(r) approximately 5.4 kda) from pyrococcus furiosus. couplings from cadmium to backbone amide protons that are hydrogen bonded to the cys-s atoms directly bonded to cd vary from smaller than 0.3 to 1.8 h ... | 1992 | 1422158 |
| glutamate dehydrogenase from the hyperthermophile pyrococcus furiosus. thermal denaturation and activation. | pyrococcus furiosus is a marine hyperthermophile that grows optimally at 100 degrees c. glutamate dehydrogenase (gdh) from p. furiosus is a hexamer of identical subunits and has an m(r) = 270,000 +/- 5500 at 25 degrees c. electron micrographs showed that the subunit arrangement is similar to that of gdh from bovine liver (i.e. 3/2 symmetry in the form of a triangular antiprism). however, gdh from p. furiosus is inactive at temperatures below 40 degrees c and undergoes heat activation above 40 de ... | 1992 | 1429615 |
| proton nmr investigation of the oxidized three-iron clusters in the ferredoxins from the hyperthermophilic archae pyrococcus furiosus and thermococcus litoralis. | the 3fe forms of ferredoxins (fd) from the hyperthermophilic archaebacteria pyrococcus furiosus (pf) and thermococcus litoralis (tl) have been investigated by 1h nmr. a combination of one-dimensional nuclear overhauser and two-dimensional noesy and bond correlation spectroscopy provides the assignment of the aromatic residues, one conserved valine, and the location of the signals for each of the three cysteines coordinated to the clusters. dipolar contacts between the trp 2 and tyr 46 in pf fd a ... | 1992 | 1445925 |
| characterization of an extremely thermostable glutamate dehydrogenase: a key enzyme in the primary metabolism of the hyperthermophilic archaebacterium, pyrococcus furiosus. | glutamate dehydrogenase (l-glutamate:nad(p)+ oxidoreductase, deaminating, ec 1.4.1.3) from the hyperthermophilic archeon pyrococcus furiosus was purified to homogeneity by chromatography on anion-exchange, molecular-exclusion and hydrophobic-interaction media. the purified native enzyme had an m(r) of 270,000 +/- 15,000 and was shown to be a hexamer with identical subunits of m(r) 46,000. the enzyme was exceptionally thermostable, having a half-life of 3.5 to more than 10 h at 100 degrees c, dep ... | 1992 | 1576153 |
| the dna polymerase from the archaebacterium pyrococcus furiosus does not testify for a specific relationship between archaebacteria and eukaryotes. | | 1992 | 1579479 |
| di-myo-inositol-1,1'-phosphate: a new inositol phosphate isolated from pyrococcus woesei. | a new inositol derivative could be isolated from the archaeum pyrococcus woesei and identified as di-myo-inositol-1,1'-phosphate by 1h, 31p nmr spectroscopy, mass spectrometry and thin layer chromatography. in p. woesei, this inositol phosphate represents the dominant counterion of k+ which ranges from 500 to 600 mm. the role of the potassium salt of di-myo-inositol-1,1'-phosphate as thermostabilizer is discussed. | 1992 | 1633880 |
| potentiometric and electron nuclear double resonance properties of the two spin forms of the [4fe-4s]+ cluster in the novel ferredoxin from the hyperthermophilic archaebacterium pyrococcus furiosus. | pyrococcus furiosus ferredoxin contains a single [4fe-4s] that exists in both s = 1/2 (20%) and s = 3/2 (80%) ground states in the reduced protein. we report here on the temperature-dependent potentiometric properties of the two spin forms, their stability, and on the structural features that differentiate them. the midpoint potential (em) of the cluster in either spin state was determined at -365 mv (30 degrees c, ph 8.0). by rapidly freezing samples for epr analyses, it was shown that the em v ... | 1991 | 1655785 |
| nucleotide sequence of a dna region comprising the gene for elongation factor 1 alpha (ef-1 alpha) from the ultrathermophilic archaeote pyrococcus woesei: phylogenetic implications. | the gene encoding elongation factor 1 alpha (ef-1 alpha, 1290 bp) of the ultrathermophilic, sulfur-reducing archaeote pyrococcus woesei was localized within a bg/ii fragment of chromosomal dna. sequence analysis showed that the ef-1 alpha gene is the upstream unit of a three-gene cluster comprising the genes for ribosomal protein s10 (306 bp) and transfer rnaser (gga). the three genes follow each other immediately in the order ef-1 alpha.s10.trna(ser) after a putative promoter located 55 bp upst ... | 1991 | 1723106 |
| high-fidelity amplification using a thermostable dna polymerase isolated from pyrococcus furiosus. | a thermostable dna polymerase which possesses an associated 3'-to-5' exonuclease (proofreading) activity has been isolated from the hyperthermophilic archaebacterium, pyrococcus furiosus (pfu). to test its fidelity, we have utilized a genetic assay that directly measures dna polymerase fidelity in vitro during the polymerase chain reaction (pcr). our results indicate that pcr performed with the dna polymerase purified from p. furiosus yields amplification products containing less than 10% of the ... | 1991 | 1761218 |
| the dna polymerase gene from the hyperthermophilic marine archaebacterium, pyrococcus furiosus, shows sequence homology with alpha-like dna polymerases. | | 1991 | 1762925 |
| extremely thermostable glutamate dehydrogenase from the hyperthermophilic archaebacterium pyrococcus furiosus. | the hyperthermophilic archaebacterium pyrococcus furiosus contains high levels of nad(p)-dependent glutamate dehydrogenase activity. the enzyme could be involved in the first step of nitrogen metabolism, catalyzing the conversion of 2-oxoglutarate and ammonia to glutamate. the enzyme, purified to homogeneity, is a hexamer of 290 kda (subunit mass 48 kda). isoelectric-focusing analysis of the purified enzyme showed a pi of 4.5. the enzyme shows strict specificity for 2-oxoglutarate and l-glutamat ... | 1991 | 1765079 |
| purification and some properties of the tungsten-containing carboxylic acid reductase from clostridium formicoaceticum. | judged by properties observed during the purification and based on the sequence of the first 25 amino acids, the enzyme from clostridium formicoaceticum catalysing the reversible reduction of non-activated carboxylic acids to aldehydes at the expense of reduced viologens, is astonishingly different from that found by us in c. thermoaceticum. according to native and sds gel electrophoresis the reductase is nearly homogeneous after only 26-fold purification. the specificity for various substrates ... | 1991 | 1793519 |
| distribution of folates and modified folates in extremely thermophilic bacteria. | analyses were made of the structures and levels of folates and modified folates present in extremely thermophilic bacteria. these procedures involved the chemical analysis of products resulting from the oxidative cleavage of the 6-substituted, folatelike tetrahydropterins present in the cells. air-oxidized cell extracts of extreme thermophiles from two members of the archaebacterial order thermococcales, thermococcus celer and pyrococcus furiosus, contained only 7-methylpterin, indicating that t ... | 1991 | 1900506 |
| the novel tungsten-iron-sulfur protein of the hyperthermophilic archaebacterium, pyrococcus furiosus, is an aldehyde ferredoxin oxidoreductase. evidence for its participation in a unique glycolytic pathway. | the anaerobic archaebacterium, pyrococcus furiosus, grows optimally at 100 degrees c by a fermentative-type metabolism in which h2, co2, and organic acids are end products. the growth of this organism is stimulated by tungsten, and, from it, a novel, red-colored, tungsten-iron-sulfur protein, abbreviated rtp, has been purified (mukund, s., and adams, m. w. w. (1990) j. biol. chem. 265, 11508-11516). rtp (mr approximately 85,000) contained approximately 1w, 7fe, and 5 acid-labile sulfide atoms/mo ... | 1991 | 1907273 |
| determinants of protein hyperthermostability: purification and amino acid sequence of rubredoxin from the hyperthermophilic archaebacterium pyrococcus furiosus and secondary structure of the zinc adduct by nmr. | the purification, amino acid sequence, and two-dimensional 1h nmr results are reported for the rubredoxin (rd) from the hyperthermophilic archaebacterium pyrococcus furiosus, an organism that grows optimally at 100 degrees c. the molecular mass (5397 da), iron content (1.2 +/- 0.2 g-atom of fe/mol), uv-vis spectrophotometric properties, and amino acid sequence (60% sequence identity with clostridium pasteurianum rd) are found to be typical of this class of redox protein. however, p. furiosus rd ... | 1991 | 1932012 |
| the hyperthermophilic archaebacterium, pyrococcus furiosus. development of culturing protocols, perspectives on scaleup, and potential applications. | from this brief discussion, it is clear that there are many obstacles to overcome before hyperthermophilic archaebacteria will be an important aspect of biotechnology. nevertheless, the prospects are intriguing. the nature of the environments that harbor these organisms and the consequent requirements for their controlled culture suggest that chemical and biochemical engineers can play an important role in elucidating their scientific and technological aspects. | 1990 | 2113371 |
| characterization of sodium dodecyl sulfate-resistant proteolytic activity in the hyperthermophilic archaebacterium pyrococcus furiosus. | cell extracts from pyrococcus furiosus were found to contain five proteases, two of which (s66 and s102) are resistant to sodium dodecyl sulfate (sds) denaturation. cell extracts incubated at 98 degrees c in the presence of 1% sds for 24 h exhibited substantial cellular proteolysis such that only four proteins could be visualized by amido black-coomassie brilliant blue staining of sds-polyacrylamide gels. the sds-treated extract retained 19% of the initial proteolytic activity as represented by ... | 1990 | 2117873 |
| spectroscopic characterization of the novel iron-sulfur cluster in pyrococcus furiosus ferredoxin. | pyrococcus furiosus ferredoxin is the only known example of a ferredoxin containing a single [4fe-4s] cluster that has non-cysteinyl ligation of one iron atom, as evidenced by the replacement of a ligating cysteine residue by an aspartic acid residue in the amino acid sequence. the properties of the iron-sulfur cluster in both the aerobically and anaerobically isolated ferredoxin have been characterized by epr, magnetic circular dichroism, and resonance raman spectroscopies. the anaerobically is ... | 1990 | 2160461 |
| purification and characterization of an alpha-glucosidase from a hyperthermophilic archaebacterium, pyrococcus furiosus, exhibiting a temperature optimum of 105 to 115 degrees c. | pyrococcus furiosus is a strictly anaerobic hyperthermophilic archaebacterium with an optimal growth temperature of about 100 degrees c. when this organism was grown in the presence of certain complex carbohydrates, the production of several amylolytic enzymes was noted. these enzymes included an alpha-glucosidase that was located in the cell cytoplasm. this alpha-glucosidase has been purified 310-fold and corresponded to a protein band of 125 kilodaltons as resolved by 10% sodium dodecyl sulfat ... | 1990 | 2163383 |
| characterization of a tungsten-iron-sulfur protein exhibiting novel spectroscopic and redox properties from the hyperthermophilic archaebacterium pyrococcus furiosus. | the archaebacterium, pyrococcus furiosus, is a strict anaerobe that grows optimally at 100 degrees c by a fermentative-type metabolism in which h2 and co2 are the only detectable products. tungsten is known to stimulate the growth of this organism. a red-colored tungsten-containing protein (abbreviated rtp) that is redox-active and extremely thermostable has been purified. rtp is a monomer of mr = 85,000 and contains approximately 6 iron, 1 tungsten, and 4 acid-labile sulfide atoms/molecule. tit ... | 1990 | 2164004 |
| glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaebacterium pyrococcus woesei: characterization of the enzyme, cloning and sequencing of the gene, and expression in escherichia coli. | the glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaebacterium pyrococcus woesei (optimal growth temperature, 100 to 103 degrees c) was purified to homogeneity. this enzyme was strictly phosphate dependent, utilized either nad+ or nadp+, and was insensitive to pentalenolactone like the enzyme from the methanogenic archaebacterium methanothermus fervidus. the enzyme exhibited a considerable thermostability, with a 44-min half-life at 100 degrees c. the amino acid sequence ... | 1990 | 2165475 |
| phylogenetic conservation of antigenic determinants in archaebacterial elongation factors (tu proteins). | by using affinity chromatography methods, we have purified elongation factor tu (ef-tu) proteins from a host of archaebacteria covering all known divisions in the archaebacterial tree except halophiles, and from such distantly related eubacteria as thermotoga maritima and escherichia coli. polyclonal antibodies were raised against the tu proteins of sulfolobus solfataricus, thermoproteus tenax, thermococcus celer, pyrococcus wosei, archaeoglobus fulgidus, methanococcus thermolitotrophicus, therm ... | 1989 | 2470483 |
| nucleotide sequence of the 5s ribosomal rna of the archaebacterium pyrococcus woesei. | | 1989 | 2503816 |
| characterization of hydrogenase from the hyperthermophilic archaebacterium, pyrococcus furiosus. | the archaebacterium, pyrococcus furiosus, grows optimally at 100 degrees c by a fermentative type metabolism in which h2 and co2 are the only detectable products. the organism also reduces elemental sulfur (s0) to h2s. cells grown in the absence of s0 contain a single hydrogenase, located in the cytoplasm, which has been purified 350-fold to apparent homogeneity. the yield of h2 evolution activity from reduced methyl viologen at 80 degrees c was 40%. the hydrogenase has a mr value of 185,000 +/- ... | 1989 | 2538471 |
| a novel and remarkably thermostable ferredoxin from the hyperthermophilic archaebacterium pyrococcus furiosus. | the archaebacterium pyrococcus furiosus is a strict anaerobe that grows optimally at 100 degrees c by a fermentative-type metabolism in which h2 and co2 are the only detectable products. a ferredoxin, which functions as the electron donor to the hydrogenase of this organism was purified under anaerobic reducing conditions. it had a molecular weight of approximately 12,000 and contained 8 iron atoms and 8 cysteine residues/mol but lacked histidine or arginine residues. reduction and oxidation of ... | 1989 | 2542225 |
| the role of posttranscriptional modification in stabilization of transfer rna from hyperthermophiles. | the influence of posttranscriptional modification on structural stabilization of trna from hyperthermophilic archaea was studied, using pyrococcus furiosus (growth optimum 100 degrees c) as a primary model. optical melting temperatures (tm) of unfractionated trna in 20 mm mg2+ are 97 degrees c for p. furiosus and 101.5 degrees c for pyrodictium occultum (growth optimum, 105 degrees c). these values are approximately 20 degrees c higher than predicted solely from g-c content and are attributed pr ... | 1994 | 7516708 |
| isolation of maltose-regulated genes from the hyperthermophilic archaeum, pyrococcus furiosus, by subtractive hybridization. | the hyperthermophilic archaeum, pyrococcus furiosus, utilizes maltose as a preferred carbon source for growth. 32p-labeled complementary dna (cdna) probes representing maltose-regulated genes were obtained by a subtractive hybridization procedure that minimized retrieval of ribosomal rna (rrna) sequences during screening. genomic dna clones were isolated by positive hybridization to these probes. genes whose expression varied both in the level of transcription, relative to rrna, as well as in th ... | 1994 | 7523251 |
| 1h nmr investigation of the electronic and molecular structure of the four-iron cluster ferredoxin from the hyperthermophile pyrococcus furiosus. identification of asp 14 as a cluster ligand in each of the four redox states. | the molecular and electronic structure of the four-iron cluster of the ferredoxin (fd) from the hyperthermophilic archaeon, pyrococcus furiosus, pf (which has only three cys in the cluster binding consensus sequence), has been investigated by 1h nmr in order to determine the identity of the noncysteinyl cluster ligand in each of the four redox states [gorst, c. m., zhou, z. h., ma, k., teng, q., howard, j. b., adams, m. w., & la mar, g. n. (1995) biochemistry 34, 8788-8795], and to characterize ... | 1995 | 7547865 |
| molecular dynamics simulations of rubredoxin from clostridium pasteurianum: changes in structure and electrostatic potential during redox reactions. | molecular dynamics simulations of clostridium pasteurianum rubredoxin in the oxidized and reduced forms have been performed. good agreement between both forms and crystal data has been obtained (rms deviation of backbone atoms of 1.06 and 1.42 a, respectively), which was due in part to the use of explicit solvent and counterions. the reduced form exhibits an unexpected structural change: the redox site becomes much more solvent-accessible, so that water enters a channel between the surface and t ... | 1995 | 7567963 |
| isolation and characterization of a heat-stable pullulanase from the hyperthermophilic archaeon pyrococcus woesei after cloning and expression of its gene in escherichia coli. | the gene encoding an extremely heat-stable pullulanase from the hyperthermophilic archaeon pyrococcus woesei was cloned and expressed in escherichia coli. purification of the enzyme to homogeneity was achieved after heat treatment of the recombinant e. coli cells, affinity chromatography on a maltotriose-coupled sepharose 6b column, and anion-exchange chromatography on mono q. the pullulanase, which was purified 90-fold with a final yield of 15%, is composed of a single polypeptide chain with a ... | 1995 | 7574598 |
| dimeric 3-phosphoglycerate kinases from hyperthermophilic archaea. cloning, sequencing and expression of the 3-phosphoglycerate kinase gene of pyrococcus woesei in escherichia coli and characterization of the protein. structural and functional comparison with the 3-phosphoglycerate kinase of methanothermus fervidus. | the gene coding for the 3-phosphoglycerate kinase (ec 2.7.2.3) of pyrococcus woesei was cloned and sequenced. the gene sequence comprises 1230 bp coding for a polypeptide with the theoretical m(r) of 46,195. the deduced protein sequence exhibits a high similarity (46.1% and 46.6% identity) to the other known archaeal 3-phosphoglycerate kinases of methanobacterium bryantii and methanothermus fervidus [fabry, s., heppner, p., dietmaier, w. & hensel, r. (1990) gene 91, 19-25]. by comparing the 3-ph ... | 1995 | 7588750 |
| aspartyl-trna synthetase of the hyperthermophilic archaeon pyrococcus sp. kod1 has a chimerical structure of eukaryotic and bacterial enzymes. | the aspartyl-trna synthetase (asprs)-encoding gene from the archaeon, pyrococcus sp. kod1 (kod1), was cloned and sequenced, and expressed in escherichia coli. the purified asprs possessed an aminoacyation activity for trna extracted from kod1. analysis of the deduced amino-acid sequence (438 aa, 50,893 da) revealed that the asprs of kod1 is a chimerical protein of bacteria and eukarya. regional analysis showed high sequence similarity to higher eukaryotic enzymes in the central and c-terminal re ... | 1995 | 7590306 |
| extreme resistance to thermally induced dna backbone breaks in the hyperthermophilic archaeon pyrococcus furiosus. | pyrococcus furiosus is a hyperthermophilic archaeon that grows optimally at 100 degrees c. it is not conceivable that these organisms could survive with genomic dna that was subject to thermal destruction, yet the mechanisms protecting the genomes of this and other hyperthermophiles against such destruction are obscure. we have determined the effect of elevated temperatures up to 110 degrees c on the molecular weight of dna in intact p. furiosus cells, compared with the effect of elevated temper ... | 1995 | 7592404 |
| molecular cloning of the transcription factor tfiib homolog from sulfolobus shibatae. | the archaea (archaebacteria) constitute a group of prokaryotes that are phylogenetically distinct from eucarya (eukaryotes) and bacteria (eubacteria). although archaea possess only one rna polymerase, evidence suggests that their transcriptional apparatus is similar to that of eucarya. for example, archaea contain a homolog of the tata-binding protein which interacts with the tata-box like a-box sequence upstream of many archaeal genes. here, we report the cloning of a sulfolobus shibatae gene t ... | 1995 | 7597084 |
| participation of the disulfide bridge in the redox cycle of the ferredoxin from the hyperthermophile pyrococcus furiosus: 1h nuclear magnetic resonance time resolution of the four redox states at ambient temperature. | the oxidized and reduced forms of the [4fe-4s]-containing ferredoxin from the hyperthermophilic archaeon pyrococcus furiosus, pf, have been investigated by 1h nuclear magnetic resonance spectroscopy, electron paramagnetic resonance spectroscopy and thiol titrations. we have identified and isolated at ambient temperature four distinct redox states for the [4fe-4s] form of the ferredoxin. these states differ in the redox state of the cluster, which is coordinated by cys 11, asp 14, cys 17, and cys ... | 1995 | 7612619 |
| redox properties of the sulfhydrogenase from pyrococcus furiosus. | the sulfhydrogenase from the extreme thermophile pyrococcus furiosus has been re-investigated. the alpha beta gamma delta heterotetrameric enzyme of 153.3 kda was found to contain 17 fe, 17 s2-, and 0.74 ni. the specific activity of the purified protein was 80 u/mg. three epr signals were found. a rhombic s = 1/2 signal (g = 2.07, 1.93, 1.89) was observed reminiscent in its shape and temperature dependence of spectra from [4fe-4s](2+; 1+) clusters. however, in reductive titrations the spectrum a ... | 1995 | 7615063 |
| cloning and sequencing of a gene from the archaeon pyrococcus furiosus with high homology to a gene encoding phosphoenolpyruvate synthetase from escherichia coli. | a gene from the hyperthermophilic archaeon pyrococcus furiosus, strain vc1 (dsm 3638), contains an 817-amino-acid open reading frame which shows 42% identity to the phosphoenolpyruvate (pep) synthetase of escherichia coli. this putative p. furiosus pep synthetase is slightly larger than the e. coli enzyme, the region between residues 58 and 89 being absent from the latter. | 1995 | 7628701 |
| response of rubredoxin from pyrococcus furiosus to environmental changes: implications for the origin of hyperthermostability. | the bases of the hyperthermostability of rubredoxin from pyrococcus furiosus (rdpf) have been probed by structural perturbations induced by solution ph and ionic strength changes. comparison of the solution behavior at ph 7 and ph 2, as probed by far- and near-uv circular dichroism, trp fluorescence emission, 1-anilinonaphthalene-8-sulfonate (ans) binding, and nmr spectroscopy, reveals the presence of only minimal structural variations at room temperature. at ph 2, the protein displays a surpris ... | 1995 | 7632687 |
| purification, characterization, and metabolic function of tungsten-containing aldehyde ferredoxin oxidoreductase from the hyperthermophilic and proteolytic archaeon thermococcus strain es-1. | thermococcus strain es-1 is a strictly anaerobic, hyperthermophilic archaeon that grows at temperatures up to 91 degrees c by the fermentation of peptides. it is obligately dependent upon elemental sulfur (s(o)) for growth, which it reduces to h2s. cell extracts contain high aldehyde oxidation activity with viologen dyes as electron acceptors. the enzyme responsible, which we term aldehyde ferredoxin oxidoreductase (aor), has been purified to electrophoretic homogeneity. aor is a homodimeric pro ... | 1995 | 7642503 |
| molecular characterization of the genes encoding the tungsten-containing aldehyde ferredoxin oxidoreductase from pyrococcus furiosus and formaldehyde ferredoxin oxidoreductase from thermococcus litoralis. | the hyperthermophilic archaea pyrococcus furiosus and thermococcus litoralis contain the tungstoenzymes aldehyde ferredoxin oxidoreductase, a homodimer, and formaldehyde ferredoxin oxidoreductase, a homotetramer. herein we report the cloning and sequencing of the p. furiosus gene aor (605 residues; m(r), 66,630) and the t. litoralis gene for (621 residues; m(r), 68,941). | 1995 | 7642512 |
| protein splicing: characterization of the aminosuccinimide residue at the carboxyl terminus of the excised intervening sequence. | protein splicing is a self-catalyzed, posttranslational process which converts a precursor polypeptide into two new proteins by the excision of an internal polypeptide segment and the ligation of the flanking polypeptides. evidence has been presented that protein splicing involves a branched intermediate, which is resolved into the two protein products by the cyclization of an asparagine residue to aminosuccinimide [xu, m. q., comb, d. g., paulus, h., noren, c. j., shao, y., & perler, f. (1994) ... | 1995 | 7662664 |
| mutational analysis using denaturing gradient gel electrophoresis and pcr. | denaturing gradient gel electrophoresis (dgge) separates (dna) molecules based on their sequence. using the proper conditions, all base-pair substitutions can be resolved from the wild-type sequence using dgge. polymerase chain reaction (pcr) permits rapid amplification of a given region of the genome. in this paper, we demonstrate the utility of dgge combined with pcr for mutation analysis by presenting different examples: (i) analysis of mouse p53 cdna for mutations, (ii) simultaneous analysis ... | 1993 | 7686254 |
| regulation of ribosomal rna transcription by growth rate of the hyperthermophilic archaeon, pyrococcus furiosus. | we have studied the single rrna gene cluster from the archaeon, pyrococcus furiosus. this isolate grows optimally at 100 degrees c and is thus a hyperthermophile. in p. furiosus, transcription of 16s rrna is subject to regulation over a 7.5-fold range in response to a 20-fold increase in growth rate. the single cluster encoding the 16s and 23s rrna genes of p. furiosus was cloned and the 1.9 kb region upstream of the 16s rrna gene was sequenced. | 1993 | 7691680 |
| characterization of the locus encoding the [ni-fe] sulfhydrogenase from the archaeon pyrococcus furiosus: evidence for a relationship to bacterial sulfite reductases. | the hydbgda genes, which encode the four subunits beta, gamma, delta and alpha of the [ni-fe] hydrogenase from the archaeon pyrococcus furiosus, have been isolated and sequenced using a pcr/ipcr-based strategy. from the sequence analysis it appears that the four structural genes are tightly linked and organized in a single transcription unit. the hydd and hyda gene products are related to the small and the large subunits of several archaeal and eubacterial [ni-fe] hydrogenases with an overall de ... | 1995 | 7704275 |
| root of the universal tree of life based on ancient aminoacyl-trna synthetase gene duplications. | universal trees based on sequences of single gene homologs cannot be rooted. iwabe et al. [iwabe, n., kuma, k.-i., hasegawa, m., osawa, s. & miyata, t. (1989) proc. natl. acad. sci. usa 86, 9355-9359] circumvented this problem by using ancient gene duplications that predated the last common ancestor of all living things. their separate, reciprocally rooted gene trees for elongation factors and atpase subunits showed bacteria (eubacteria) as branching first from the universal tree with archaea (a ... | 1995 | 7708661 |
| pyruvate: ferredoxin oxidoreductase from the sulfate-reducing archaeoglobus fulgidus: molecular composition, catalytic properties, and sequence alignments. | archaeoglobus fulgidus is a hyperthermophilic sulfate-reducing archaeon. in this communication we describe the purification and properties of pyruvate: ferredoxin oxidoreductase from this organism. the catabolic enzyme was purified 250-fold to apparent homogeneity with a yield of 16%. the native enzyme had an apparent molecular mass of 120 kda and was composed of four different subunits of apparent molecular masses of 45, 33, 25, and 13 kda, indicating an alpha beta gamma delta structure. per mo ... | 1995 | 7710318 |
| glyceraldehyde-3-phosphate ferredoxin oxidoreductase, a novel tungsten-containing enzyme with a potential glycolytic role in the hyperthermophilic archaeon pyrococcus furiosus. | the archaeon pyrococcus furiosus grows optimally at 100 degrees c by the fermentation of carbohydrates to yield acetate, co2, and h2. cell-free extracts contain very low activity of the glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase, but extremely high activity of glyceraldehyde-3-phosphate ferredoxin oxidoreductase (gapor). gapor was purified under strictly anaerobic conditions. it is a monomeric, o2-sensitive protein of m(r) approximately 63,000 which contains pterin and approxima ... | 1995 | 7721730 |
| 1h nuclear-magnetic-resonance investigation of oxidized fe4s4 ferredoxin from thermotoga maritima. hyperfine-shifted resonances, sequence-specific assignments and secondary structure. | the oxidized fe4s4 ferredoxin from the hyperthermophilic bacterium thermotoga maritima has been investigated by one- and two-dimensional nmr in order to characterize its hyperfine-shifted resonances originating from the cysteinyl cluster ligands and to assign its resonances in the diamagnetic shift range. the chemical shift and relaxation time pattern of the hyperfine-shifted signals is very similar to other oxidized fe4s4 ferredoxins. a tentative sequence-specific assignment of these resonances ... | 1995 | 7758460 |
| insights into thermal stability from a comparison of the glutamate dehydrogenases from pyrococcus furiosus and thermococcus litoralis. | in the light of the solution of the three-dimensional structure of the nad(+)-linked glutamate dehydrogenase from the mesophile clostridium symbiosum, we have undertaken a detailed examination of the alignment of the sequences for the thermophilic glutamate dehydrogenases from thermococcus litoralis and pyrococcus furiosus against the sequence and the molecular structure of the glutamate dehydrogenase from c. symbiosum, to provide insights into the molecular basis of their thermostability. this ... | 1995 | 7758464 |
| purification and properties of extremely thermostable glutamate dehydrogenases from two hyperthermophilic archaebacteria, pyrococcus woesei and pyrococcus furiosus. | glutamate dehydrogenase (l-glutamate: nadp oxidoreductase, deaminating, ec 1.4.1.4) from the hyperthermophilic archaebacteria pyrococcus woesei and p. furiosus were purified to homogeneity from crude extracts. the enzymes had similar enzymological properties: molecular mass, subunit structure, optimum phs for the oxidative deamination and reductive amination, substrate specificity and coenzyme specificity as well as thermostability; the activity was not lost after incubation at 105 degrees c for ... | 1993 | 7763882 |
| a variable-temperature direct electrochemical study of metalloproteins from hyperthermophilic microorganisms involved in hydrogen production from pyruvate. | the hyperthermophilic bacterium thermotoga maritima and the hyperthermophilic archaeon pyrococcus furiosus grow optimally at 80 and 100 degrees c, respectively, by the fermentation of carbohydrates to organic acids, co2, and h2. pyruvate is a major source of reductant for h2 production during fermentation, and pyruvate ferredoxin oxidoreductase (por), a 4fe-type ferredoxin, and hydrogenase have been previously purified from both species. p. furiosus utilizes a copper-iron-containing por and a ni ... | 1995 | 7766626 |
| the eubacterial transcriptional activator lrp is present in the archaeon pyrococcus furiosus. | | 1995 | 7770911 |
| rapid differentiation of hyperthermophilic archaea by restriction mapping of the intergenic spacer regions of the ribosomal rna operons. | we report a sensitive and convenient method for rapid differentiation of new isolates of hyperthermophilic archaea. polymerase chain reaction (pcr) was used to amplify the intergenic spacer regions of the ribosomal rna operons of eight archaea. spacer regions from one euryarcheote, pyrococcus furiosus, and one crenarcheote, pyrodictium brockii, were sequenced completely. restriction fragment length polymorphism (rflp) analyses were performed on the spacer regions from eight hyperthermophilic arc ... | 1995 | 7773330 |
| purification and characterization of a thermostable thiol protease from a newly isolated hyperthermophilic pyrococcus sp. | a hyperthermophilic archaeon strain, kod1, was isolated from a solfatara at a wharf on kodakara island, kagoshima, japan. the growth temperature of the strain ranged from 65 to 100 degrees c, and the optimal temperature was 95 degrees c. the anaerobic strain was an s0-dependent heterotroph. cells were irregular cocci and were highly motile with several polar flagella. the membrane lipid was of the ether type, and the gc content of the dna was estimated to be 38 mol%. the 16s rrna sequence was 95 ... | 1994 | 7811092 |
| 1h nmr investigation of the paramagnetic cluster environment in pyrococcus furiosus three-iron ferredoxin: sequence-specific assignment of ligated cysteines independent of tertiary structure. | one- and two-dimensional 1h nmr data tailored to detect paramagnetically relaxed protons near the s = 1/2, three-iron-sulfur cluster of the ferredoxin from the hyperthermophile pyrococcus furiosus are analyzed to sequence specifically assign the hyperfine shifted ligated cysteine signals, to determine the nature of the secondary structural elements on which these cysteines reside, and to define the tertiary contacts of the cluster with the remainder of the previously characterized secondary stru ... | 1995 | 7819255 |
| isolation, sequence and characterization of the maltose-regulated mlra gene from the hyperthermophilic archaeum pyrococcus furiosus. | the mlra (maltose regulated) gene from the hyperthermophilic archaeum pyrococcus furiosus was identified from a family of clones whose expression was influenced by the presence of maltose in the medium. sequencing of the 2276 bp of dna containing mlra and flanking regions revealed a 753-amino-acid (aa) (88 kda) open reading frame (orf). the orf is preceded by a bacterial-like ribosome-binding site. the deduced product shared extensive homology with pyruvate dikinases (pdk) from both eukaryal and ... | 1994 | 7828869 |
| a gene from the hyperthermophile pyrococcus furiosus whose deduced product is homologous to members of the prolyl oligopeptidase family of proteases. | the mlr-2 gene from the hyperthermophilic archaeum pyrococcus furiosus was identified from a family of clones whose expression was influenced by the presence of maltose in the medium. the sequence of 2100 bp of dna containing mlr-2 and its flanking regions revealed a 616-amino-acid (71 kda) open reading frame (orf). the orf's initiation codon appeared 10 nt into the mlr-2 message and was not preceded by any apparent ribosome-binding site. the deduced product shared homology with prolyl endopepti ... | 1995 | 7828913 |
| an extremely thermostable aromatic aminotransferase from the hyperthermophilic archaeon pyrococcus furiosus. | pyrococcus furiosus is a strictly anaerobic archaeon (formerly archaebacterium) that grows optimally at 100 degrees c by the fermentation of peptides. cell-free extracts were found to contain two distinct aromatic aminotransferases (arat, ec 2.6.1.57), one of which was purified to electrophoretic homogeneity. p. furiosus arat is a homodimer with a subunit m(r) value of 44,000 +/- 1000. using 2-ketoglutarate as the amino acceptor, the purified enzyme catalyzed the pyridoxal 5'-phosphate (pmp)-dep ... | 1995 | 7873596 |
| structure of a hyperthermophilic tungstopterin enzyme, aldehyde ferredoxin oxidoreductase. | the crystal structure of the tungsten-containing aldehyde ferredoxin oxidoreductase (aor) from pyrococcus furiosus, a hyperthermophilic archaeon (formerly archaebacterium) that grows optimally at 100 degrees c, has been determined at 2.3 angstrom resolution by means of multiple isomorphous replacement and multiple crystal form averaging. aor consists of two identical subunits, each containing an fe4s4 cluster and a molybdopterin-based tungsten cofactor that is analogous to the molybdenum cofacto ... | 1995 | 7878465 |
| expression and in vitro assembly of recombinant glutamate dehydrogenase from the hyperthermophilic archaeon pyrococcus furiosus. | the gdha gene, encoding the hexameric glutamate dehydrogenase (gdh) from the hyperthermophilic archaeon pyrococcus furiosus, was expressed in escherichia coli by using the pet11-d system. the recombinant gdh was soluble and constituted 15% of the e. coli cell extract. the n-terminal amino acid sequence of the recombinant protein was identical to the sequence of the p. furiosus enzyme, except for the presence of an initial methionine which was absent from the enzyme purified from p. furiosus. by ... | 1995 | 7887598 |
| the purification, cloning, and high level expression of a glutaredoxin-like protein from the hyperthermophilic archaeon pyrococcus furiosus. | a protein has been purified to homogeneity from crude extracts of the hyperthermophilic archaeon pyrococcus furiosus based on its ability to catalyze the reduction of insulin disulfides in the presence of dithiothreitol; the protein has a molecular mass of 24.8 kda and a pi of 4.9, and it is highly heat-stable. the first 29 amino acid residues at the n terminus of the p. furiosus protein were determined by edman degradation, and its gene was cloned in escherichia coli. the amino acid sequence de ... | 1995 | 7890703 |
| chromosomal organization and nucleotide sequence of the fus-gene encoding elongation factor 2 (ef-2) of the hyperthermophilic archaeum pyrococcus woesei. | a pyrococcus woesei ecori dna fragment (3400 bp) harbouring the gene fus for elongation factor 2 (ef-2) was cloned and almost completely sequenced. unlike methanococcus vannielii (which displays the 'str operon'-like fus and tuf gene context, 5'-rps12-rps7-fus-tuf-3'), and similar to sulfolobus acidocaldarius and desulfurococcus mobilis, the pyrococcus fus gene (732 codons) is unlinked to the rps and tuf genes, and is immediately followed (57 bp intergenic spacing) by an orf of 106 codons. both ... | 1995 | 7896082 |
| chromosomal organization and nucleotide sequence of the genes for elongation factors ef-1 alpha and ef-2 and ribosomal proteins s7 and s10 of the hyperthermophilic archaeum desulfurococcus mobilis. | the desulfurococcus mobilis genes fus (encoding ef-2) and tuf (for ef-1 alpha) were cloned and sequenced together with genes for ribosomal proteins s10 (rps10) and s7 (rps7). unlike methanococcus, which displays the bacterial-like fus and tuf gene context 5'-rps12-rps7-fus-tuf-3', and similar to sulfolobus and pyrococcus, the desulfurococcus fus gene (734 codons) has a distinct chromosomal location. moreover, tuf (441 codons) is the promoter-proximal unit of a three-gene cluster comprising the g ... | 1995 | 7898436 |
| evolutionary relationships of bacterial and archaeal glutamine synthetase genes. | glutamine synthetase (gs), an essential enzyme in ammonia assimilation and glutamine biosynthesis, has three distinctive types: gsi, gsii and gsiii. genes for gsi have been found only in bacteria (eubacteria) and archaea (archaebacteria), while gsii genes only occur in eukaryotes and a few soil-dwelling bacteria. gsiii genes have been found in only a few bacterial species. recently, it has been suggested that several lateral gene transfers of archaeal gsi genes to bacteria may have occurred. in ... | 1994 | 7916055 |
| robust amplification and ethidium-visible detection of the fragile x syndrome cgg repeat using pfu polymerase. | we report on a method for ethidium bromide detection of fmr1 normal and premutation-size cgg triplet repeats. a set of pcr conditions has been optimized for the polymerase of the hyperthermophilic bacterium, pyrococcus furiosus. using this protocol, normal-size alleles ranging from 5 to 50 repeats, as well as most of premutation-size alleles, varying from > 50 to approximately 200 repeats, could be detected by ethidium bromide staining of agarose gels. since the protocol requires neither autorad ... | 1994 | 7943034 |
| biochemical diversity among sulfur-dependent, hyperthermophilic microorganisms. | hyperthermophiles are a recently discovered group of microorganisms that grow at and above 90 degrees c. they currently comprise over 20 different genera, and except for two novel bacteria, all are classified as archaea. the majority of these organisms are obligately anaerobic heterotrophs that reduce elemental sulfur (s degree) to h2s. the best studied from a biochemical perspective are the archaeon, pyrococcus furiosus, and the bacterium, thermotoga maritima, both of which are saccharolytic. p ... | 1994 | 7946471 |
| histone-encoding genes from pyrococcus: evidence for members of the hmf family of archaeal histones in a non-methanogenic archaeon. | two genes, designated hpya1 and hpya2, have been cloned and sequenced from pyrococcus strain gb-3a. they are predicted to encode proteins (hpya1 and hpya2, respectively) that are approx. 60% identical to the histones hmf and hmt, characterized from methanogenic archaea. these archaeal histones also contain the amino-acid sequences, conserved in eukaryotic h4 histones, that are thought to interact directly with dna. | 1994 | 7959058 |
| sulfide dehydrogenase from the hyperthermophilic archaeon pyrococcus furiosus: a new multifunctional enzyme involved in the reduction of elemental sulfur. | pyrococcus furiosus is an anaerobic archaeon that grows optimally at 100 degrees c by the fermentation of carbohydrates yielding acetate, co2, and h2 as the primary products. if elemental sulfur (s0) or polysulfide is added to the growth medium, h2s is also produced. the cytoplasmic hydrogenase of p. furiosus, which is responsible for h2 production with ferredoxin as the electron donor, has been shown to also catalyze the reduction of polysulfide to h2s (k. ma, r. n. schicho, r. m. kelly, and m. ... | 1994 | 7961401 |
| occurrence of tertiary and quaternary branched polyamines in thermophilic archaebacteria. | polyamines of thermophilic archaebacteria were analysed by high-performance liquid chromatography and gas chromatography. thermoplasma acidophilum and thermoplasma volcanium ubiquitously contained spermidine and spermine. four species of sulfolobus, s. acidocaldarius, s. solfataricus, s. metallicus and s. shibatae, two species of acidianus, a. brierleyi and a. infernus, and metallosphaera sedula, contained norspermidine and norspermine in addition to spermidine and spermine, but quantitative dis ... | 1994 | 7968661 |
| evidence for the operation of a novel embden-meyerhof pathway that involves adp-dependent kinases during sugar fermentation by pyrococcus furiosus. | the main pathway for the fermentation of maltose or cellobiose by the hyperthermophile pyrococcus furiosus was investigated by in vivo nmr and by enzyme measurements. addition of [1-13c]glucose to cell suspensions resulted in the formation of c2-labeled acetate and c3-labeled alanine. no label was recovered in co2 or hco3-. in the presence of [3-13c]glucose, the label ended up in the c1 atom of alanine and in hco3- and co2. these labeling patterns indicate that glucose is converted along an embd ... | 1994 | 8021261 |
| cloning, expression, and molecular characterization of the gene encoding an extremely thermostable [4fe-4s] ferredoxin from the hyperthermophilic archaeon pyrococcus furiosus. | the gene for ferredoxin from the hyperthermophilic archaeon pyrococcus furiosus was cloned, sequenced, and expressed in escherichia coli. the coding region confirmed the determined amino acid sequence. putative archaeon-type transcriptional regulatory elements were identified. the fdxa gene appears to be an independent transcriptional unit. recombinant ferredoxin was indistinguishable from the protein purified from p. furiosus in its thermal stability and in the potentiometric and spectroscopic ... | 1994 | 8045914 |
| the amino acid sequence of glutamate dehydrogenase from pyrococcus furiosus, a hyperthermophilic archaebacterium. | the complete amino acid sequence of glutamate dehydrogenase from the archaebacterium pyrococcus furiosus has been determined. the sequence was reconstructed by automated sequence analysis of peptides obtained after cleavage with cyanogen bromide, asp-n endoproteinase, trypsin, or pepsin. the enzyme subunit is composed of 420 amino acid residues yielding a molecular mass of 47,122 d. in the recently determined primary structure of glutamate dehydrogenase from another thermophilic archaebacterium, ... | 1994 | 8060497 |
| improved efficiency in site-directed mutagenesis by pcr using a pyrococcus sp. gb-d polymerase. | | 1994 | 8068335 |
| the hyperthermophilic glycolytic enzyme enolase in the archaeon, pyrococcus furiosus: comparison with mesophilic enolases. | high enolase activity, as measured by the conversion of 2-phosphoglycerate to phosphoenolpyruvate, was found in the cytoplasm of pyrococcus furiosus (an anaerobic, hyperthermophilic archaeon that grows optimally at 100 degrees c). in this organism, the enzyme probably functions in a sugar fermentation pathway. the enzyme was purified to homogeneity. it had a temperature optimum of > 90 degrees c and a ph optimum of 8.1. the enzyme was extremely thermostable with a time for 50% inactivation at 10 ... | 1994 | 8080274 |
| growth requirements of hyperthermophilic sulfur-dependent heterotrophic archaea isolated from a shallow submarine geothermal system with reference to their essential amino acids. | three hyperthermophilic sulfur-dependent heterotrophs were isolated from a shallow submarine hydrothermal system at an inlet of kodakara-jima island, kagoshima, japan. the isolates grew at 60 to 97 degrees c, with the optimum temperatures at 85 to 90 degrees c. sensitivity to rifampin and the existence of ether lipids indicated that the isolates are hyperthermophilic archaea. partial sequencing of the genes coding for 16s rrna showed that the three isolates are closely related to the genus therm ... | 1994 | 8085828 |
| cloning and sequencing of the gene encoding glutamine synthetase i from the archaeum pyrococcus woesei: anomalous phylogenies inferred from analysis of archaeal and bacterial glutamine synthetase i sequences. | the gene glna encoding glutamine synthetase i (gsi) from the archaeum pyrococcus woesei was cloned and sequenced with the sulfolobus solfataricus glna gene as the probe. an operon reading frame of 448 amino acids was identified within a dna segment of 1,528 bp. the encoded protein was 49% identical with the gsi of methanococcus voltae and exhibited conserved regions characteristic of the gsi family. the p. woesei gsi was aligned with available homologs from other archaea (s. solfataricus, m. vol ... | 1993 | 8098326 |
| evolution of translational elongation factor (ef) sequences: reliability of global phylogenies inferred from ef-1 alpha(tu) and ef-2(g) proteins. | the ef-2 coding genes of the archaea pyrococcus woesei and desulfurococcus mobilis were cloned and sequenced. global phylogenies were inferred by alternative tree-making methods from available ef-2(g) sequence data and contrasted with phylogenies constructed from the more conserved but shorter ef-1 alpha(tu) sequences. both the monophyly (sensu hennig) of archaea and their subdivision into the kingdoms crenarchaeota and euryarchaeota are consistently inferred by analysis of ef-2(g) sequences, us ... | 1994 | 8159735 |
| sequence, assembly and evolution of a primordial ferredoxin from thermotoga maritima. | a gene coding for the ferredoxin of the primordial, strictly anaerobic and hyperthermophilic bacterium thermotoga maritima was cloned, sequenced and expressed in escherichia coli. the ferredoxin gene encodes a polypeptide of 60 amino acids that incorporates a single 4fe-4s cluster. t. maritima ferredoxin expressed in e. coli is a heat-stable, monomeric protein, the spectroscopic properties of which show that its 4fe-4s cluster is correctly assembled within the mesophilic host, and that it remain ... | 1994 | 8168477 |
| secretion of thermostable dna polymerase using a novel baculovirus vector. | baculovirus-mediated expression has become a powerful tool for the high yield production of functionally active recombinant proteins. in order to further enhance the utility of this expression system, we constructed versatile transfer vectors that facilitate the secretion of recombinant proteins from host insect cells by inserting functional secretory leader sequences down-stream of the polyhedrin promoter. in-frame insertion of cdna sequences results in the synthesis of fusion proteins containi ... | 1994 | 8175786 |
| transcription factor iid in the archaea: sequences in the thermococcus celer genome would encode a product closely related to the tata-binding protein of eukaryotes. | the first step in transcription initiation in eukaryotes is mediated by the tata-binding protein, a subunit of the transcription factor iid complex. we have cloned and sequenced the gene for a presumptive homolog of this eukaryotic protein from thermococcus celer, a member of the archaea (formerly archaebacteria). the protein encoded by the archaeal gene is a tandem repeat of a conserved domain, corresponding to the repeated domain in its eukaryotic counterparts. molecular phylogenetic analyses ... | 1994 | 8183889 |
| the tata-binding protein: a general transcription factor in eukaryotes and archaebacteria. | the tata-binding protein tbp appears to be essential for all transcription in eukaryotic cell nuclei, which suggests that its function was established early in evolution. archaebacteria constitute a kingdom of organisms distinct from eukaryotes and eubacteria. archaebacterial gene regulatory sequences often map to tata box-like motifs. here it is shown that the archaebacterium pyrococcus woesei expresses a protein with structural and functional similarity to eukaryotic tbp molecules. this sugges ... | 1994 | 8191287 |
| solution 1h nmr determination of secondary structure for the three-iron form of ferredoxin from the hyperthermophilic archaeon pyrococcus furiosus. | two-dimensional 1h nmr data have been used to make sequence-specific assignments and define the secondary structure of the three-iron form of the oxidized ferredoxin, fd, from the hyperthermophilic archaeon pyrococcus furiosus, pf. signals for at least some protons were located for 65 of the 66 amino acids in the sequence, in spite of the paramagnetic (s = 1/2) ground state, but not all could be assigned. unassigned and missing signals could be qualitatively correlated with the expected proximit ... | 1994 | 8193147 |
| indolepyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon pyrococcus furiosus. a new enzyme involved in peptide fermentation. | pyrococcus furiosus is a strictly anaerobic archaeon that grows optimally at 100 degrees c by a fermentative-type metabolism in which complex peptide mixtures such as yeast extract and tryptone, and also certain sugars, are oxidized to organic acids, h2 and co2. enzymes involved in the utilization of peptides such as proteases, aromatic amino transferases, and glutamate dehydrogenase have been previously purified from this organism. it is shown here that p. furiosus also contains significant cyt ... | 1994 | 8206994 |
| the purification and characterization of an extremely thermostable alpha-amylase from the hyperthermophilic archaebacterium pyrococcus furiosus. | the alpha-amylase from pyrococcus furiosus, a hyperthermophilic archaebacterium, has been purified to homogeneity. the enzyme is a homodimer with a subunit molecular mass of 66 kda. the isoelectric point is 4.3. the enzyme displays optimal activity, with substantial thermal stability, at 100 degrees c, with the onset of activity at approximately 40 degrees c. unlike mesophilic alpha-amylases there is no dependence on ca2+ for activity or thermostability. the enzyme displays a broad range of subs ... | 1993 | 8226989 |
| alpha-amylase from the hyperthermophilic archaebacterium pyrococcus furiosus. cloning and sequencing of the gene and expression in escherichia coli. | a gene encoding a highly thermostable alpha-amylase from the hyperthermophilic archaebacterium pyrococcus furiosus was cloned and expressed in escherichia coli. the nucleotide sequence of the gene predicts a 649-amino acid protein with a calculated molecular mass of 76.3 kda, which corresponds well with the value obtained from purified enzyme using denaturing polyacrylamide gel electrophoresis. the nh2 terminus of the deduced amino acid sequence corresponds precisely to that obtained from the pu ... | 1993 | 8226990 |
| enzymes and proteins from organisms that grow near and above 100 degrees c. | microorganisms that can grow at and above 100 degrees c were discovered a decade ago, and about 20 different genera are now known. these so-called hyperthermophiles are the most ancient of all extant life; all but two genera are classified as archaea. all have been isolated from geothermal heated environments including deep-sea hydrothermal vents. this group includes some methanogenic and sulfate-reducing species, but the majority are strictly anaerobic heterotrophs that utilize complex peptide ... | 1993 | 8257111 |
| in vitro protein splicing of purified precursor and the identification of a branched intermediate. | protein splicing is a posttranslational processing event in which an internal polypeptide is excised from a protein precursor and the terminal polypeptides are then ligated together, resulting in the production of two proteins. this report presents direct evidence for protein splicing by demonstrating in vitro splicing of purified precursor that accumulated when the protein splicing element from pyrococcus dna polymerase was cloned into a foreign gene. in vitro splicing was temperature and ph de ... | 1993 | 8269515 |
| pyruvate ferredoxin oxidoreductases of the hyperthermophilic archaeon, pyrococcus furiosus, and the hyperthermophilic bacterium, thermotoga maritima, have different catalytic mechanisms. | pyruvate ferredoxin oxidoreductase (por) has been previously purified from two hyperthermophiles, the archaeon pyrococcus furiosus (pf, topt = 100 degrees c) and the bacterium thermotoga maritima (tm, topt = 80 degrees c). each catalyzes the oxidative decarboxylation of pyruvate to acetyl-coa and co2 near the optimal growth temperature of the organism and are virtually inactive at 25 degrees c. both pors contain a thiamine pyrophosphate (tpp) cofactor and at least two [4fe-4s] ferredoxin-type cl ... | 1994 | 8305427 |
| low incorporation of dump by some thermostable dna polymerases may limit their use in pcr amplifications. | incorporation of dump instead of dtmp is frequently used to control carryover contamination during pcr amplifications. we have tested four thermostable dna polymerases for their ability to utilize dutp as a substrate in pcr. amplification of products in the presence of dutp instead of dttp was good with thermus aquaticus dna polymerase but highly inefficient with three other thermostable dna polymerases. the latter was due to: (a) lower incorporation of dump relative to dtmp, (b) increased proof ... | 1993 | 8323030 |
| complete nucleotide sequence of an archaeal (pyrococcus woesei) gene encoding a homolog of eukaryotic transcription factor iib (tfiib). | | 1993 | 8332505 |
| characterization, cloning, and in vitro expression of the extremely thermostable glutamate dehydrogenase from the hyperthermophilic archaeon, es4. | glutamate dehydrogenase (gdh) from the hyperthermophilic archaeon es4 (optimal growth temperature 98 degrees c and maximum growth temperature 110 degrees c) was purified to homogeneity. the purified native enzyme had an m(r) of 270,000 +/- 5,000 and was shown by gel filtration and sds-polyacrylamide gel electrophoresis to be a hexamer with identical subunits of m(r) = 46,000 +/- 3,000. the hexameric subunit composition was also evident from electron micrographs, which show a triangular antiprism ... | 1993 | 8349661 |
| [pyroglycolysis, a novel glycolytic pathway in pyrococcus furiosus]. | | 1993 | 8372233 |
| purification and characterization of pyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon pyrococcus furiosus. | pyrococcus furiosus grows optimally at 100 degrees c by carbohydrate fermentation. it is thought to contain a novel tungsten-dependent, nad(p)-independent glycolytic pathway in which one of the oxidation steps is catalyzed by a tungsten-containing aldehyde ferredoxin oxidoreductase. the enzyme that catalyzes the terminal oxidation step, pyruvate ferredoxin oxidoreductase (por), has now been purified. por has a molecular mass of 100 kda and is comprised of three subunits (45, 31 and 24 kda). it l ... | 1993 | 8380721 |
| hydrogenase of the hyperthermophile pyrococcus furiosus is an elemental sulfur reductase or sulfhydrogenase: evidence for a sulfur-reducing hydrogenase ancestor. | microorganisms growing near and above 100 degrees c have recently been discovered near shallow and deep sea hydrothermal vents. most are obligately dependent upon the reduction of elemental sulfur (s0) to hydrogen sulfide (h2s) for optimal growth, even though s0 reduction readily occurs abiotically at their growth temperatures. the sulfur reductase activity of the anaerobic archaeon pyrococcus furiosus, which grows optimally at 100 degrees c by a metabolism that produces h2s if s0 is present, wa ... | 1993 | 8389482 |