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a seven-iron ferredoxin from the thermoacidophilic archaeon desulfurolobus ambivalens.a seven-iron ferredoxin was isolated from aerobically grown cells of the hyperthermoacidophilic archaeon desulfurolobus ambivalens (dsm 3772). the protein is monomeric, with an apparent molecular mass of 15 kda and contains 7 iron atoms/molecule. the n-terminal sequence shows a large similarity (70% identity) with that of the ferredoxin isolated from the archaeon sulfolobus acidocaldarius. the epr characteristics in both the native (oxidized) and dithionite-reduced states of this protein allowed ...19957851403
the thermosome of thermoplasma acidophilum and its relationship to the eukaryotic chaperonin tric.a high molecular-mass protein complex from the archaebacterium thermoplasma acidophilum, referred to here as the 'thermosome', is built from two subunits (m(r) 58 and 60). the thermosome has been purified to homogeneity. the molecular mass of the native complex was determined to be 1061 +/- 30 da by scanning transmission electron microscopy. it shows a weak atpase activity and is able to bind denatured polypeptides. averages obtained from electron micrographs of negatively stained molecules in t ...19957867646
expression of a synthetic gene encoding p2 ribonuclease from the extreme thermoacidophilic archaebacterium sulfolobus solfataricus in mesophylic hosts.this work reports the molecular cloning and expression of a synthetic gene encoding p2, a 7-kda ribonuclease (rnase) previously isolated in our laboratory from the archaebacterium sulfolobus solfataricus [fusi et al., eur. j. biochem. 211 (1993) 305-310]. the p2-encoding synthetic gene was expressed in e. coli and in saccharomyces cerevisiae. the recombinant (re-) protein was produced to approx. 1.5% of the total protein content in s. cerevisiae using the galactose-inducible gal1 promoter and to ...19957867957
cloning and functional analysis of the tata binding protein from sulfolobus shibatae.archaea (formerly archaebacteria) comprise a domain of life that is phylogenetically distinct from both eucarya and bacteria. here we report the cloning of a gene from the archaeon sulfolobus shibatae that encodes a protein with strong homology to the tata binding protein (tbp) of eukaryotes. sulfolobus shibatae tbp is, however, almost as diverged from other archaeal tbps that have been cloned as it is from eukaryotic tbps. dna binding studies indicate that s.shibatae tbp recognizes tata-like a- ...19957784182
primary structure of the thermosome from thermoplasma acidophilum.the thermosome, a chaperonin from the archaebacterium thermoplasma acidophilum, consists of two subunits (m(r) 58,000 and 60,000) which assemble into a cylindrical complex of pseudo eight-fold rotational symmetry. the sequences of the two subunits are approximately 60% identical to each other and to tf55 from sulfolobus shibatae, and are 30-40% identical to the subunits of the tcp1 containing ring complex (tric) from the eukaryotic cytosol. a dendrogram of this family of chaperonins contains eig ...19957794526
molecular cloning, nucleotide sequence, and expression of a carboxypeptidase-encoding gene from the archaebacterium sulfolobus solfataricus.mammalian metallocarboxypeptidases play key roles in major biological processes, such as digestive-protein degradation and specific proteolytic processing. a sulfolobus solfataricus gene (cpsa) encoding a recently described zinc carboxypeptidase with an unusually broad substrate specificity was cloned, sequenced, and expressed in escherichia coli. despite the lack of overall sequence homology with known carboxypeptidases, seven homology blocks, including the zn-coordinating and catalytic residue ...19957559343
conformational cycle of the archaeosome, a tcp1-like chaperonin from sulfolobus shibatae.the major heat shock proteins in the archaeon sulfolobus shibatae are similar to the cytosolic eukaryotic chaperonin and form an 18-subunit bitoroidal complex. two sequence-related subunits constitute a functional complex, named the archaeosome. the archaeosome exists in two distinct conformational states that are part of chaperonin functional cycle. the closed archaeosome complex binds atp and forms an open complex. upon atp hydrolysis, the open complex dissociates into subunits. free subunits ...19957499406
thermopsin. 19957674919
the first nucleotide sequence of an archaeal elongation factor 1 beta gene.an archaeal elongation factor 1 beta gene has been isolated for the first time from a sulfolobus solfataricus genomic library. the sequenced clone (869 bp) contained two open reading frames, one coding for a protein made of 91 amino acid residues (ssef-1 beta), the other one encoding a nonidentified product (orf 115). the amino acid sequences of segments at the n- and c-terminal of the translated ssef-1 beta were identical to those determined for the native protein. northern and southern analyse ...19957632739
purification and characterization of the rieske iron-sulfur protein from the thermoacidophilic crenarchaeon sulfolobus acidocaldarius.the previously detected rieske iron-sulfur protein from the membranes of the thermoacidophile sulfolobus acidocaldarius [anemüller, s., et al. (1993) febs lett. 318, 61-64] was purified to electrophoretic homogeneity and the n-terminal amino acids determined. the apparent molecular weight was estimated to be 32 kda. the reduced protein displays a rhombic epr spectrum with gxyz = 1.768, 1.895, 2.035. the average g-value of 1.902 is typical for nitrogen ligand-containing clusters. epr spin quantif ...19957867807
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 ...19957898436
stability against temperature and external agents of vesicles composed of archael bolaform lipids and egg pc.the bolaform lipid ple extracted from the thermophilic archaeon sulfolobus solfataricus and its mixtures with egg phosphatidyl-choline (egg pc) have been used to prepare sonicated vesicles. the leakage of entrapped calcein was continuously monitored by fluorescence dequenching. the half times of leakage have been used to compare vesicle stability under different conditions of temperature, lipid composition and presence of destabilizing agents like ca2+ ions and poly(ethylene glycol) (peg). it ha ...19957495852
a secy homologous gene in the crenarchaeon sulfolobus acidocaldarius.the nucleotide sequence of an open reading frame, located upstream of the gene for adenylate kinase, was determined in the thermoacidophile crenarchaeon sulfolobus acidocaldarius. data bank searches identified the sequence as a secy homologous gene. the dna derived protein sequence of total 463 amino acids contains 10 hydrophobic domains. a sequence alignment with other prokaryotic and eukaryotic secy sequences reveals significant homology, but the secy primary sequence of s. acidocaldarius show ...19957495856
in the thermophilic archaeon sulfolobus solfataricus a dna-binding protein is in vitro (adpribosyl)ated.a small protein with high affinity for homologous dna was isolated from sulfolobus solfataricus homogenate by mineral acid extraction. it was purified using a two-step procedure including cm-cellulose and rp-hpl chromatographies. the protein was electrophoretically homogeneous, had a molecular weight of 7.147 kda and an amino acid composition with a high content of lysine and glutamic acid residues. the protein was able to protect dna against thermal denaturation and dnase i digestion in a dose- ...19957887965
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 ...19957896082
alternative form of the dicluster ferredoxin from the thermoacidophilic archaeon, sulfolobus sp. strain 7.an alternative form of a 7fe dicluster-type ferredoxin (fd-b) with a different charge density was purified as a minor component from the aerobic and thermoacidophilic archaeon, sulfolobus sp. strain 7. comparison of its properties with those of native 7fe ferredoxin (fd-a), a major ferredoxin, was made in terms of the molecular properties, the absorption, circular dichroism and electron paramagnetic resonance spectral properties, and the reactivity coupled with the cognate 2-oxoacid:ferredoxin o ...19957826373
an 8.5-kda ribonuclease from the extreme thermophilic archaebacterium sulfolobus solfataricus.protein p3, a ribonuclease we previously isolated from the archaebacterium sulfolobus solfataricus [p. fusi et al. (1993) eur. j. biochem. 211, 305-310], was subjected to complete amino acid sequencing. it consisted of 75 residues, with a calculated m(r) of 8582, a pi of 10.1, and had some degree of monomethylation at lys-4 and lys-6. p2, a previously sequenced, 62-residue ribonuclease from the same organism, had an identical sequence for 57 consecutive residues starting from the n-terminus. p2 ...19957875327
glyceraldehyde-3-phosphate dehydrogenase in the hyperthermophilic archaeon sulfolobus solfataricus: characterization and significance in glucose metabolism.glyceraldehyde-3-phosphate dehydrogenase in the archaeon sulfolobus solfataricus (ssgapd) has been purified 232 fold with an overall recovery of about 25%. the enzyme is a homomeric tetramer with an m(r) of 41 kda/subunit. it utilizes either nad+ or nadp+ as coenzyme but its affinity for the latter is about 50 fold higher. ssgapd activity is maximum at 87 degrees c. in the range 45-87 degrees c the arrhenius plot is linear and the activation energy is 55 kj/mol. the enzyme is thermostable, with ...19957663406
molecular properties of glutamate dehydrogenase from the extreme thermophilic archaebacterium sulfolobus solfataricus.this study is concerned with the structural characterization in solution of the glutamate dehydrogenase from the archaeon sulfolobus solfataricus. at neutral ph both alpha-helix and beta-sheet constitute the secondary structure of this enzyme, on the basis of circular dichroism. a complex, temperature dependent self-association equilibrium regulates the formation of the enzyme quaternary structure, which seems to be accompanied by a reversible structural change. at 25 degrees c the enzyme is mos ...19957669806
a gene encoding a putative membrane protein homologous to the major facilitator superfamily of transporters maps upstream of the beta-glycosidase gene in the archaeon sulfolobus solfataricus.we have identified a gene encoding a putative membrane protein homologous to the major facilitator superfamily, mapping upstream of the lacs gene in sulfolobus solfataricus. permeases from this family mediate secondary transport and are widely distributed among eubacteria and eukaryotes; the finding of an archaeal member suggests that this mechanism of transport evolved before the divergence of the three living domains. we also report a transcriptional mapping of the gene cluster.19957533760
purification and characterization of 3-isopropylmalate dehydrogenase from a thermoacidophilic archaebacterium sulfolobus sp. strain 7.3-isopropylmalate dehydrogenase was purified (about 2000-fold) to homogeneity for the first time from an archaebacterium, sulfolobus sp. strain 7. the enzyme showed an apparent molecular mass of about 110 kda by gel filtration and a single 36-kda polypeptide band on sds-page, suggesting tri- or tetrameric structure. the pi value was 6.9. the n-terminal amino acid sequence was similar to enzymes from other sources. the enzyme activity was greatly stimulated by the presence of mn2+, cd2+, mg2+, or ...19957557336
expression and extensive characterization of a beta-glycosidase from the extreme thermoacidophilic archaeon sulfolobus solfataricus in escherichia coli: authenticity of the recombinant enzyme.the gene coding for the beta-glycosidase from the archaeon sulfolobus solfataricus has been overexpressed in escherichia coli. the enzyme was purified to homogeneity with a rapid purification procedure employing a thermal precipitation as a crucial step. the final yield was 64% and the purification from the thermal precipitation was 5.4-fold. the expressed enzyme shows the same molecular mass, thermophilicity, thermal stability, and broad substrate specificity, with noticeable exocellobiase (glu ...19958541022
expression of an archaeal chaperonin in e. coli: formation of homo- (alpha, beta) and hetero-oligomeric (alpha+beta) thermosome complexes.co-expression of the two genes encoding the alpha- and beta-subunit of the thermoplasma acidophilum thermosome in escherichia coli yielded fully assembled hetero-oligomeric complexes (alpha+beta). surprisingly, also separate expression of both genes resulted in formation of hexadecameric complexes (alpha, beta) in the bacterial cytoplasm. on electron micrographs these complexes were indistinguishable from each other and from the native thermosome. the recombinant alpha-complex as well as the nat ...19958521970
resolution of the aerobic respiratory system of the thermoacidophilic archaeon, sulfolobus sp. strain 7. i. the archaeal terminal oxidase supercomplex is a functional fusion of respiratory complexes iii and iv with no c-type cytochromes.the aerobic respiratory system of the thermoacidophilic archaeon, sulfolobus sp. strain 7, is unusual in that it consists of only a- and b-type cytochromes but no c-type cytochromes. in previous studies, a novel cytochrome oxidase a583-aa3 subcomplex has been purified, which showed a ferrocytochrome c oxidase but no caldariellaquinol oxidase activity (wakagi, t., yamauchi, t., oshima, t., müller, m., azzi, a., and sone, n. (1989) biochem. biophys. res. commun. 165, 1110-1114). we show here that ...19958537342
resolution of the aerobic respiratory system of the thermoacidophilic archaeon, sulfolobus sp. strain 7. ii. characterization of the archaeal terminal oxidase subcomplexes and implication for the intramolecular electron transfer.the terminal segment of the aerobic respiratory chain of the thermoacidophilic archaeon sulfolobus sp. strain 7 is an unusual caldariellaquinol oxidase supercomplex, which contains at least one b-type and three spectroscopically distinguishable a-type cytochromes, one copper, and a rieske-type fes center. in this paper, we report the purification and characterization of two different forms of the archaeal a-type cytochromes, namely, a three-subunit cytochrome a583-aa3 subcomplex and a single-sub ...19958537343
identification of the iron-sulfur clusters in a ferredoxin from the archaeon sulfolobus acidocaldarius. evidence for a reduced [3fe-4s] cluster with ph-dependent electronic properties.a ferredoxin isolated from the archaeon sulfolobus acidocaldarius strain dsm 639 has been shown to contain one [3fe-4s]1 + 10 cluster with a reduction potential of -275 mv and one [4fe-4s]2+/1+ cluster with a reduction potential of -529 mv at ph 6.4, in the temperature range 0-50 degrees c. the monomer molecular mass was confirmed to be 10907.5 +/- 1.0 da by electrospray mass spectrometry, as calculated from the published amino acid sequence [minami, y. wakabayashi. s., wada, k., matsubara, h., ...19958521862
the phosphoglycerate kinase and glyceraldehyde-3-phosphate dehydrogenase genes from the thermophilic archaeon sulfolobus solfataricus overlap by 8-bp. isolation, sequencing of the genes and expression in escherichia coli.the overlapping genes encoding phosphoglycerate kinase (pgk) and glyceraldehyde-3-phosphate dehydrogenase (grap-dh) from the hyperthermophilic archaeon sulfolobus solfataricus have been cloned and sequenced. pcr primers based on highly conserved regions of different pgk sequences were used to isolate an internal region of the pgk gene. this was then used to screen a genomic library to isolate the full length pgk gene. a 2.5-kb bglii fragment of s. solfataricus dna contained both the pgk gene and ...19958521845
industrial-scale production and rapid purification of an archaeal beta-glycosidase expressed in saccharomyces cerevisiae.the application of enzymes isolated from extreme thermophiles in biotechnological processes is hampered by their unconventional fermentation conditions. the expression, in mesophilic hosts, of genes encoding for thermophilic proteins enables these difficulties to be overcome and permits the production of enzymes in high yield by using conventional fermentation plants and an efficient enzyme purification utilizing heat precipitation of host proteins. the beta-glycosidase gene from sulfolobus solf ...19958573288
structure-based sequence alignment of elongation factors tu and g with related gtpases involved in translation.the g domain and domain ii in the crystal structure of thermus thermophilus elongation factor g (ef-g) were compared with the homologous domains in thermus aquaticus elongation factor tu (ef-tu). sequence alignment derived from the structural superposition was used to define conserved sequence elements in domain ii. these elements and previously known conserved sequence elements in the g domain were used to guide the alignment of the sequences of sulfolobus acidocaldarius elongation factor 2, hu ...19958587108
isocitrate dehydrogenases from haloferax volcanii and sulfolobus solfataricus: enzyme purification, characterisation and n-terminal sequence.the isocitrate dehydrogenases from the extremely halophilic archaeon, haloferax volcanii, and from the hyperthermophilic archaeon, sulfolobus solfataricus, have been purified to electrophoretic homogeneity. the purified enzymes have been characterised with respect to their cofactor specificities, subunit compositions and their salt and thermal stabilities. n-terminal amino acid sequences have been determined for both enzymes, and multiple alignments with sequences of bacterial and eukaryotic iso ...19958593959
how to make my blood boil.two recent papers comparing the structure of a hyperthermophilic protein with its mesophilic counterpart both conclude that large networks of ion-pairs are important for hyperthermostability. how and why is not yet clear.19958747452
a potential anti-oxidant protein in a ferrous iron-oxidizing sulfolobus species.the production of a 25 kda protein was greatly increased when a sulfur- and ferrous iron-oxidizing species of sulfolobus was switched from growth on tetrathionate to growth on ferrous iron. the gene encoding the protein was cloned and sequenced. the predicted amino sequence showed significant similarity to those of the alkyl hydroperoxide reductase/thiol specific anti-oxidant family of proteins that appear to be involved in responses to certain types of oxidative stress.19958593961
resolution of the aerobic respiratory system of the thermoacidophilic archaeon, sulfolobus sp. strain 7. iii. the archaeal novel respiratory complex ii (succinate:caldariellaquinone oxidoreductase complex) inherently lacks heme group.an active respiratory complex ii (succinate:quinone oxidoreductase) has been purified from tetraether lipid membranes of the thermoacidophilic archaeon, sulfolobus sp. strain 7. it consists of four different subunits with apparent molecular masses of 66, 37, 33, and 12 kda on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. the 66-kda subunit contains a covalently bound flavin, the 37-kda subunit is a possible iron-sulfur protein carrying three distinct types of epr-visible fes cluster ...19958537344
nucleic acid binding properties of a helix stabilising nucleoid protein from the thermoacidophilic archaeon sulfolobus acidocaldarius that condenses dna into compact structures.helix stabilising nucleoid protein (hsnp-c') from an acidothermophilic archaeon sulfolobus acidocaldarius has been characterised with respect to interaction with nucleic acids by gel retardation assay, binding to nucleic acid columns, fluorescence titrations and electron microscopy. the protein exists in solution as very large multimeric aggregates as indicated by cross-linking studies. the protein binds strongly and co-operatively to double stranded dna. electron microscopy of the complexes of ...19958714217
preribosomal rna processing in archaea: characterization of the rnp endonuclease mediated processing of precursor 16s rrna in the thermoacidophile sulfolobus acidocaldarius.the hyperthermophilic archaeon sulfolobus acidocaldarius uses a novel rna-containing endonuclease to excise and mature 16s rrna from the precursor (pre) rrna transcript. a cell-free processing system has been developed using an in vitro transcribed rna substrate containing the entire 144 nucleotide 5' external transcribed spacer (5'ets) and the first 72 nucleotides of 16s rrna. the cell-free extract cleaves in the 5'ets at positions -99, -31, and +1 (i.e., the 5'ets-16s junction). these position ...19958721997
2.0 a structure of indole-3-glycerol phosphate synthase from the hyperthermophile sulfolobus solfataricus: possible determinants of protein stability.recent efforts to understand the basis of protein stability have focused attention on comparative studies of proteins from hyperthermophilic and mesophilic organisms. most work to date has been on either oligomeric enzymes or monomers comprising more than one domain. such studies are hampered by the need to distinguish between stabilizing interactions acting between subunits or domains from those acting within domains. in order to simplify the search for determinants of protein stability we have ...19958747456
intercellular mobility and homing of an archaeal rdna intron confers a selective advantage over intron- cells of sulfolobus acidocaldarius.some intron-containing rrna genes of archaea encode homing-type endonucleases, which facilitate intron insertion at homologous sites in intron- alleles. these archaeal rrna genes, in contrast to their eukaryotic counterparts, are present in single copies per cell, which precludes intron homing within one cell. however, given the highly conserved nature of the sequences flanking the intron, homing may occur in intron- rrna genes of other archaeal cells. to test whether this occurs, the intron-con ...19958618886
on the origin of respiration: electron transport proteins from archaea to man.all aerobic organisms use the exergonic reduction of molecular oxygen to water as primary source of metabolic energy. this reaction is catalyzed by membrane residing terminal heme/cu-oxidases which belong to a superfamily of widely varying structural complexity between mitochondrial and bacterial members of this family. over the last few years, considerable information from this and other laboratories accumulated also on archaeal respiratory chains and their terminal oxidases. in the following, ...19968639327
methionine aminopeptidase-1: the map of the mitochondrion? 19968772380
viruses, plasmids and other genetic elements of thermophilic and hyperthermophilic archaea.we review and update the work on genetic elements, e.g., viruses and plasmids (exluding is elements and transposons) in the kingdom crenarchaeota (thermoproteales and sulfolobales) and the orders thermococcales and thermoplasmales in the kingdom euryarchaeota of the archael domain, including unpublished data from our laboratory. the viruses of crenarchaeota represent four novel virus families. the fuselloviridae represented by ssvi of s. shibatae and relatives in other sulfolobus strains have th ...19968639330
the unique dna topology and dna topoisomerases of hyperthermophilic archaea.hyperthermophilic archaea exhibit a unique pattern of dna topoisomerase activities. they have a peculiar enzyme, reverse gyrase, which introduces positive superturns into dna at the expense of atp. this enzyme has been found in all hyperthermophiles tested so far (including bacteria) but never in mesophiles. reverse gyrases are formed by the association of a helicase-like domain and a 5'-type 1 dna topoisomerase. these two domains might be located on the same polypeptide. however, in the methano ...19968639331
general vectors for archaeal hyperthermophiles: strategies based on a mobile intron and a plasmid.although there are currently no cloning and expression vectors available for archaeal hyperthermophiles, small cryptic plasmids have been characterized for these organisms as well as viruses and introns capable of spreading between cells. below, we review the recent progress in adapting these genetic elements as vectors for pyrococcus furiosus and sulfolobus acidocaldarius. an efficient and reliable transformation procedure is described for both organisms. the potential of the mobile intron from ...19968639332
the pressure-dependence of two beta-glucosidases with respect to their thermostability.a comparative study of temperature and pressure effects were carried out by using two homologous enzymes exhibiting different thermostability and oligomery: almond beta-glucosidase and sulfolobus solfataricus beta-glucosidase. both the activity and stability were studied using an in-house built bioreactor allowing injection, stirring, sampling and on-line spectrophometric monitoring with retention of pressure up to 2.5 kbar and temperature control possible up to 150 degrees c. almond beta-glucos ...19968645739
archaeal elongation factor 1 beta is a dimer. primary structure, molecular and biochemical properties.the elongation factor 1 beta (ef-1 beta), that in eukarya and archaea promotes the replacement of gdp by gtp on the elongation factor 1 alpha x gdp complex, was purified to homogeneity from the thermoacidophilic archaeon sulfolobus solfataricus (ssef-1 beta). its primary structure was established by sequenced edman degradation of the entire protein or its proteolytic peptides. the molecular weight of ssef-1 beta was estimated as about 10000 or 20000 under denaturing or native conditions respecti ...19968652615
thermal unfolding of the dna-binding protein sso7d from the hyperthermophile sulfolobus solfataricus.thermal unfolding of the small hyperthermophilic dna-binding protein sso7d was studied by circular dichroism spectroscopy and differential scanning calorimetry. the unfolding transition can be described by a reversible two state process. maximum stability was observed in the region between ph 4.5 and 7.0 where sso7d unfolds with a melting temperature between 370.8 to 371.9 k and an unfolding enthalpy between 62.9 and 65.4 kcal/mol. the heat capacity differences between the native and the heat de ...19969000635
identification of two glutamic acid residues essential for catalysis in the beta-glycosidase from the thermoacidophilic archaeon sulfolobus solfataricus.the sulfolobus solfataricus, strain mt4, beta-glycosidase (ss beta-gly) is a thermophilic member of glycohydrolase family 1. to identify active-site residues, glutamic acids 206 and 387 have been changed to isosteric glutamine by site-directed mutagenesis. mutant proteins have been purified to homogeneity using the schistosoma japonicum glutathione s-transferase (gst) fusion system. the proteolytic cleavage of the chimeric protein with thrombin was only obtainable after the introduction of a mol ...19969010932
purification and characterization of thermostable maltooligosyl trehalose trehalohydrolase from the thermoacidophilic archaebacterium sulfolobus acidocaldarius.a thermostable maltooligosyl trehalose trehalohydrolase from the thermoacidophilic archaebacterium sulfolobus acidocaldarius atcc 33909 was purified from a cell-free extract to an electrophoretically pure state by successive column chromatographies on sepabeads fp-da13, butyl-toyopearl 650m, deae-toyopearl 650s, toyopearl hw-55s and ultrogel aca44. the enzyme had a molecular mass of 59,000 by sds-polyacrylamide gel electrophoresis and a pi of 6.1 by gel isoelectrofocusing. the n-terminal amino a ...19969063974
purification and characterization of thermostable maltooligosyl trehalose synthase from the thermoacidophilic archaebacterium sulfolobus acidocaldarius.a thermostable maltooligosyl trehalose synthase was purified from a cell-free extract of the thermoacidophilic archaebacterium sulfolobus acidocaldarius atcc 33909 to an electrophoretically homogeneous state by successive column chromatography on sepabeads fp-da13, butyl-toyopearl 650m, deae-toyopearl 650s, ultrogel aca44, and mono q. the enzyme had a molecular mass of 74,000 by sds-polyacrylamide gel electrophoresis and a pi of 5.9 by gel isoelectrofocusing. the n-terminal amino acid of the enz ...19969063973
immobilization on chitosan of a thermophilic beta-glycosidase expressed in saccharomyces cerevisiae.a sulfolobus solfataricus beta-glycosidase expressed in saccharomyces cerevisiae (s beta gly) was immobilized on chitosan activated with glutaraldehyde. the yield of immobilization was evaluated as 80%. compared to the free beta-glycosidase, the immobilized enzyme showed a similar ph optimum (ph = 7.0), the same increasing activity up to 80 degrees c, improved thermostability, and no inhibition by glucose. functional studies pointed out that the kinetic constant values for both enzymes were comp ...19969100353
the site for gtp hydrolysis on the archaeal elongation factor 2 is unmasked by aliphatic alcohols.an appropriate mixture of ethylene glycol and bacl2 enhanced the otherwise very low intrinsic gtpase activity of the elongation factor 2 isolated from the archaeon sulfolobus solfataricus (ssef-2). the enzymatic activity became up to 300-fold higher than that of the ssef-2 gtpase measured in the absence of any stimulator, but remained 20-fold lower than that stimulated by ribosome. the stimulatory effect of ethylene glycol/ba2+ was attributed to the increased affinity for gtp, probably related t ...19969116052
phylogenetic analysis of carbamoylphosphate synthetase genes: complex evolutionary history includes an internal duplication within a gene which can root the tree of life.carbamoylphosphate synthetase (cps) catalyzes the first committed step in pyrimidine biosynthesis, arginine biosynthesis, or the urea cycle. organisms may contain either one generalized or two specific cps enzymes, and these enzymes may be heterodimeric (encoded by linked or unlinked genes), monomeric, or part of a multifunctional protein. in order to help elucidate the evolution of cps, we have performed a comprehensive phylogenetic analysis using the 21 available complete cps sequences, includ ...19968752005
molecular cloning, sequencing, and heterologous expression of a novel zinc-containing ferredoxin gene from a thermoacidophilic archaeon sulfolobus sp. strain 7.the gene encoding a novel zinc-containing ferredoxin from a hyperthermophilic and acidophilic archaeon (archaebacterium) sulfolobus sp. strain 7 was cloned and sequenced. the dna sequence predicts a 103 residue protein after removal of n-terminal methionine, which is in good agreement with the results of the protein analysis. surprisingly, the residues responsible for binding a zinc atom were conserved among three other thermoacidophilic archaea. a common sequence stretch vxgxhxghx8-17pxxlgxhgtx ...19968753789
uracil phosphoribosyltransferase from the extreme thermoacidophilic archaebacterium sulfolobus shibatae is an allosteric enzyme, activated by gtp and inhibited by ctp.uracil phosphoribosyltransferase, which catalyses the formation of ump and pyrophosphate from uracil and 5-phosphoribosyl alpha-1-pyrophosphate (prpp), was partly purified from the extreme thermophilic archaebacterium sulfolobus shibatae. the enzyme required divalent metal ions for activity and it showed the highest activity at ph 6.4. the specific activity of the enzyme was 50-times higher at 95 degrees c than at 37 degrees c, but the functional half-life was short at 95 degrees c. the activity ...19968765224
a novel amidohydrolase gene from bacillus subtilis cloning: dna-sequence analysis and map position of amhx.the nucleotide sequence of a new bacillus subtilis gene (amhx) was determined that encodes a protein (amhx) with strong sequence identity to amidohydrolases from both plant and bacterial species and a carboxypeptidase from the archaeon sulfolobus sulfataricus. the amhx gene encodes a hydrophilic polypeptide of 383 amino acids with a molecular mass of 41.5 kda. the amhx gene was overexpressed in e. coli by using the t7 rna polymerase/promoter system and the transcription initiation sites for the ...19968768514
extremely thermophilic and thermostable 5'-methylthioadenosine phosphorylase from the archaeon sulfolobus solfataricus. gene cloning and amino acid sequence determination.a gene encoding an extremely thermophilic and thermostable 5'-methylthioadenosine phosphorylase was cloned from the archaeon sulfolobus solfataricus. two degenerate oligodeoxyribonucleotide probes synthesized on the basis of the n-terminal amino acid sequence of the protein were used to screen a genomic library of s. solfataricus cloned into the pgem7zf(+) vector. the dna fragment of 2118 bp containing the 5'-methylthioadenosine phosphorylase gene was sequenced. the open reading frame comprises ...19968774706
thermostable dna polymerases. 19968791630
interfacial air/water proton conduction from long distances by sulfolobus solfataricus archaeal bolaform lipids.the stability, structural organization, and the ability to transfer protons long distances have been investigated in monolayers formed from archael bolaform lipids at the air/water interface. the lipids employed were the fractions gror2gro (r represents an acyl group with variable chain length typically consisting of 0-4 cyclopentane rings and 40 isoprenoid residues) and gror2gronon-ol (non-ol represents nonitol) extracted from sulfolobus solfataricus by hydrolysis of the cytoplasmic membrane. g ...19968797852
comparison of a beta-glucosidase and a beta-mannosidase from the hyperthermophilic archaeon pyrococcus furiosus. purification, characterization, gene cloning, and sequence analysis.two distinct exo-acting, beta-specific glycosyl hydrolases were purified to homogeneity from crude cell extracts of the hyperthermophilic archaeon pyrococcus furiosus: a beta-glucosidase, corresponding to the one previously purified by kengen et al. (kengen, s. w. m., luesink, e. j., stams, a. j. m., and zehnder, a. j. b. (1993) eur. j. biochem. 213, 305-312), and a beta-mannosidase. the beta-mannosidase and beta-glucosidase genes were isolated from a genomic library by expression screening. the ...19968798600
adenylate kinase from sulfolobus acidocaldarius: expression in escherichia coli and characterization by fourier transform infrared spectroscopy.adenylate kinase from the extremely thermoacidophilic archaeon sulfolobus acidocaldarius has been overexpressed in escherichia coli. the highly purified enzyme was characterized by fourier transform infrared spectroscopy (ftir). analysis of ftir spectra and estimation of secondary structure revealed a global protein structure similar to that of other adenylate kinases. thermal unfolding of the protein with an estimated tm value near 90 degrees c is irreversible due to protein aggregation. the en ...19968806756
reverse gyrase gene from sulfolobus shibatae b12: gene structure, transcription unit and comparative sequence analysis of the two domains.we cloned and sequenced a dna fragment from the thermophilic archaeal strain sulfolobus shibatae b12 that includes the gene topr encoding the reverse gyrase. the rna of the reverse gyrase gene was characterized indicating that the topr gene is fully functional in vivo. we showed by primer extension analysis that transcription of topr initiates 28 bp downstream from a consensus a-box promoter. in order to understand how this particular type i dna topoisomerase introduces positive superturns into ...19968972852
cca-adding enzymes and poly(a) polymerases are all members of the same nucleotidyltransferase superfamily: characterization of the cca-adding enzyme from the archaeal hyperthermophile sulfolobus shibatae.we describe the purification, cloning, and characterization of the cca-adding enzyme [atp(ctp):trna nucleotidyl transferase] from the thermophilic archaebacterium, sulfolobus shibatae. characterization of an archaeal cca-adding enzyme provides formal proof that the cca-adding activity is present in all three contemporary kingdoms. antibodies raised against recombinant, expressed sulfolobus cca-adding enzyme reacted specifically with the 48-kda protein and fully depleted all cca-adding activity f ...19968809016
identification and characterization of geranylgeraniol kinase and geranylgeranyl phosphate kinase from the archaebacterium sulfolobus acidocaldarius.geranylgeranyl diphosphate is an important precursor of archaebacterial ether-linked lipids, and it has been thought that all of this compound is "de novo" synthesized by geranylgeranyl diphosphate synthase. we studied the phosphorylation of geranylgeraniol, which seems to be related to the salvage pathway of biosynthesis of archaebacterial ether-linked lipids, in the archaebacterium sulfolobus acidocaldarius. activities of geranylgeraniol kinase and geranylgeranyl phosphate kinase were detected ...19968830051
cloning and sequencing of a cluster of genes encoding novel enzymes of trehalose biosynthesis from thermophilic archaebacterium sulfolobus acidocaldarius.trehalose biosynthesis genes, trez, trex and trey, encoding maltooligosyltrehalose trehalohydrolase (trez), glycogen debranching enzyme (trex), and maltooligosyltrehalose synthase (trey) have been cloned from the thermophilic archaebacterium sulfolobus acidocaldarius atcc33909. the amino-acid sequences deduced from trez, trex and trey are composed of 556, 713 and 720 amino-acid residues, respectively. trez and trey are 33-40% homologous to the corresponding enzymes from arthrobacter sp. q36. we ...19968980629
resonance raman spectroscopy of the integral quinol oxidase complex of sulfolobus acidocaldarius.the integral quinol oxidase complex of sulfolobus acidocaldarius (dsm 639) was investigated by resonance raman spectroscopy. the complex includes four heme a groups which constitute two functional entities, a587 and aa3, containing two low-spin hemes and a low-spin as well as a high-spin heme, respectively. rr spectra were obtained from the fully oxidized and fully reduced states of the complex using different excitation wavelengths in the soret band region in order to disentangle the contributi ...19968841122
functional and structural properties of the homogeneous beta-glycosidase from the extreme thermoacidophilic archaeon sulfolobus solfataricus expressed in saccharomyces cerevisiae.a protein with beta-glycosidase activity from sulfolobus solfataricus (s beta gly) was expressed in the yeast saccharomyces cerevisiae. the purification procedure was made fast and easy by employing a single chromatographic step. after 5.8-fold purification, the cell extract gave a homogeneous enzyme at 166 u/mg. the recombinant enzyme was functionally and structurally similar to the wild-type enzyme. kinetic experiments showed the same wide substrate specificity; in fact, the expressed enzyme h ...19968860656
expression in escherichia coli of thermostable elongation factor 1 alpha from the archaeon sulfolobus solfataricus.the elongation factor 1 alpha from the archaeon sulfolobus solfataricus (ssef-1 alpha) was expressed in escherichia coli and purified. the ssef-1 alpha gene was amplified by pcr and cloned in the ndel site of the pt7-7 expression vector, under the control of the promoter of t7 rna polymerase. upon induction with isopropyl beta-d-thiogalactopyranoside, the recombinant ssef-1 alpha (recssef-1 alpha) was purified from the e. coli s-100 extract by a two-step procedure. from 1 litre of cell culture, ...19968867895
purification and characterization of nadh oxidase from the archaea sulfolobus acidocaldarius and sulfolobus solfataricus.the enzyme nadh oxidase (ec 1.6.99.3) has been isolated from the two thermoacidophilic archaea sulfolobus acidocaldarius and sulfolobus solfataricus and characterized. in both organisms the enzyme oxidizes specifically beta-nadh in the presence of molecular oxygen and requires the presence of a flavin cofactor, showing a high specificity for fad. a stoicheiometric amount of hydrogen peroxide to nadh is formed as the end product of the reaction, indicating that both enzymes are two-electron donor ...19968867896
identification of a dinb/umuc homolog in the archeon sulfolobus solfataricus.to date, eight closely related homologs of the escherichia coli umuc protein have been identified. all of these homologs appear to play critical roles in damage-inducible mutagenesis in enterobacteriaceae. recently, a distantly related umuc-homolog, dinb, has also been identified in e. coli. using the polymerase chain reaction together with degenerate primers designed against conserved regions found in umuc-like proteins, we have identified a new member of the umuc-superfamily in the archeon sul ...19968876701
the response of selected members of the archaea to the gram stain.archaea possess a broader range of cell envelope structural formats than eubacteria and their cell walls do not contain peptidoglycan. some archaea have only a single s-layer as their cell wall (e.g. methanococcus jannaschii and sulfolobus acidocaldarius), whereas others have multiple layers (e.g. methanospirillum hungatei). sometimes there can also be a high proportion of tetraether lipids in membranes to make the envelope more resilient to environmental stress (e.g. methanococcus jannaschii an ...19968885405
effect of physical constraints on the mechanisms of membrane fusion: bolaform lipid vesicles as model systems.bolaform lipid vesicles were used to study the effect of physical constraints on membrane fusion. in these vesicles the membrane is organized in a single monolayer, because of the presence of covalent bonds in its middle plane. therefore, the formation of fusion intermediates is subject to higher energy barriers and greater geometrical constraints than is usual in bilayer membranes. bolaform lipids were extracted from the thermophilic archaeon sulfolobus solfataricus. these lipids can be divided ...19968889155
temperature-induced denaturation of beta-glycosidase from the archaeon sulfolobus solfataricus.the beta-glycosidase isolated from the extreme thermophilic archaeon sulfolobus solfataricus, grown at 87 degrees c, is a tetrameric protein with a molecular mass of 240 kda. this enzyme is barely active at 30 degrees c and has optimal activity, over 95 degrees c, at ph 6.5. its thermal stability was investigated at ph 10.1 and 10.6 by means of functional studies, circular dichroism and differential scanning calorimetry. there was no evidence of thermal activation of the enzyme and the temperatu ...19968889813
the crystal structure of indole-3-glycerol phosphate synthase from the hyperthermophilic archaeon sulfolobus solfataricus in three different crystal forms: effects of ionic strength.indole-3-glycerol phosphate synthase from the hyperthermophilic archaeon sulfolobus solfataricus is a monomeric enzyme with the common (beta/alpha)8-fold. recently, its three-dimensional structure was solved in an orthorhombic crystal form, grown by using 1.3 m ammonium sulfate as precipitating agent. here we describe the x-ray structure analysis of two new crystal forms of this enzyme that were obtained at medium and low ionic strength, respectively. hexagonal crystals with space group p3(1)21 ...19968893859
organizational characteristics and information content of an archaeal genome: 156 kb of sequence from sulfolobus solfataricus p2.we have initiated a project to sequence the 3 mbp genome of the thermoacidophilic archaebacterium sulfolobus solfataricus p2. cosmids were selected from a provisional set of minimally overlapping clones, subcloned in puc18, and sequenced using a hybrid (random plus directed) strategy to give two blocks of contiguous unique sequence, respectively, 100,389 and 56,105 bp. these two contigs contain a total of 163 open reading frames (orfs) in 26-29 putative operons; 56 orfs could be identified with ...19968899719
purification and characterization of new trehalose-producing enzymes isolated from the hyperthermophilic archae, sulfolobus solfataricus km1.amylolytic activity that converts soluble starch to alpha, alpha-trehalose (trehalose), was found in the cell homogenate of the hyperthermophilic acidophilic archae, sulfolobus solfataricus km1. deae chromatography of the homogenate as well as other new reliable assay methods showed two enzymes to be essential for this activity. these enzymes, a glycosyltransferase and an amylase, were purified to homogeneity and characterized. their molecular masses were 76 kda and 61 kda and activities were ma ...19968901122
2-oxoacid:ferredoxin oxidoreductase from the thermoacidophilic archaeon, sulfolobus sp. strain 7.the purified 2-oxoacid:ferredoxin oxidoreductase of a thermoacidophilic and aerobic crenarchaeote, sulfolobus sp. strain 7, consists of 70-kda alpha and 37-kda beta subunits, and contains one thiamine pyrophosphate (tpp), one [4fe-4s]2+.1+ cluster, and two magnesium atoms per alpha beta structure. it exhibits a broad substrate specificity toward 2-oxoacids such as 2-oxoglutarate, 2-oxobutyrate, and pyruvate. the gene encoding the archaeal oxidoreductase was cloned, and the two open reading frame ...19968902625
an aspartate aminotransferase from an extremely thermophilic bacterium, thermus thermophilus hb8.the aspartate aminotransferase gene (aspat, ec 2.6.1.1) of an extremely thermophilic bacterium, thermus thermophilus hb8, was cloned and sequenced, and its gene product was overproduced. the purified t. thermophilus aspat was stable up to about 80 degrees c at neutral ph. t. thermophilus aspat was strictly specific for acidic amino acid substrates, such as aspartate, glutamate, and the respective keto acids. the gene coding for t. thermophilus aspat showed that it comprised 1,155 bp with a high ...19968907187
redox-linked ionization of sulredoxin, an archaeal rieske-type [2fe-2s] protein from sulfolobus sp. strain 7."sulredoxin" of sulfolobus sp. strain 7 is an archaeal soluble rieske-type [2fe-2s] protein and was initially characterized by several spectroscopic techniques (iwasaki, t., isogai, t., iizuka, t. , and oshima, t. (1995) j. bacteriol. 177, 2576-2582). it appears to have tightly linked ionization affecting the redox properties of the protein, which is characteristic of the rieske fes proteins found as part of the respiratory chain. sulredoxin had an em(low ph) value of +188 +/- 9 mv, and the slop ...19968910356
protein-encoding genes in the sulfothermophilic archaea sulfolobus and pyrococcus.a number of unrelated protein-encoding genes from sulfothermophilic archaea, sulfolobus acidocaldarius, sulfolobus solfataricus, pyrococcus furiosus and pyrococcus woesei, has been analyzed. in the sulfolobus genus, the content of a + t is significantly higher than that of c + g and the base usage follows the order, a > t > g > c. in pyrococcus, the a + t content is also higher than that of c + g, but with lower values; in the order of base usage, g precedes t. the codon usage of these sulfother ...19968918227
cloning and sequencing of the gene coding for s-adenosylhomocysteine hydrolase in the thermophilic archaeon sulfolobus solfataricus.the gene from the thermophilic archaeon sulfolobus solfataricus (ss), encoding the s-adenosylhomocysteine hydrolase (adohcyhd), has been cloned. two degenerate oligodeoxyribonucleotide (oligo) probes, synthesized on the basis of amino acid (aa) sequence of cyanogen bromide-peptide fragments of the purified protein, were used to screen a genomic library of ss cloned into the pgem7zf(+) vector. the adohcyhd gene (adohcyhd) comprises 1254 nucleotides (nt) and encodes a polypeptide of 417 aa with a ...19968921839
the dna polymerase-encoding gene from a thermoacidophilic archaeon sulfolobus acidocaldarius.we have cloned, sequenced and characterized the gene encoding a dna polymerase from the thermoacidophilic archaeon sulfolobus acidocaldarius (sac). the putative transcription promoter and terminator elements, as well as a potential ribosome-binding site (rbs), have been identified in the flanking regions. one large open reading frame (orf) found in the sequenced portion of the sac genome encodes a protein of 875 amino acids (aa). all conserved motifs characteristic of family b of dna polymerases ...19968921881
three extremely thermostable proteins from sulfolobus and a reappraisal of the 'traffic rules'.three cytosolic enzymes from the extremely thermoacidophilic archaeon sulfolobus acidocaldarius (dsm 639) have been investigated: adenylate kinase, pyrophosphatase and superoxide dismutase. the latter was isolated from s. acidocaldarius cells, the others were heterologically overproduced in escherichia coli. long-term thermostability, flexibility, catalytic activity, and thermal denaturation were investigated by biochemical and physical methods. superoxide dismutase is hyperthermostable over sev ...19968922285
sulfolobus hakonensis sp. nov., a novel species of acidothermophilic archaeon.we characterized a microbial strain that was isolated from a hot spring at a geothermal area in hakone, japan. this isolate, whose lobed-shaped cells were about 1.0 micron in diameter, was a facultative chemolitho-autotroph that required aerobic conditions for growth. the optimum ph was 3.0 (ph range, 1.0 to 4.0), and the optimum temperature was 70 degrees c (temperature range, 50 to 80 degrees c). lithotrophically, this strain grew on elemental sulfur and reduced sulfur compounds. the g+c conte ...19968934897
transformation of the extremely thermoacidophilic archaeon sulfolobus solfataricus via a self-spreading vector.we describe a transformation system for extremely thermophilic archaea of the genus sulfolobus in the kingdom crenarchaeota. we have constructed in vitro a recombinant derivative of the recently described conjugative plasmid pnob8, containing a beta-galactosidase gene downstream of a strong promotor. transformation of a beta-galactosidase negative mutant of sulfolobus solfataricus with this construct resulted in its spreading through the culture containing the primary transformants and in effici ...19968935654
a putative signal recognition particle receptor alpha subunit (sr alpha) homologue is expressed in the hyperthermophilic crenarchaeon sulfolobus acidocaldarius.a 1.64 kb genomic dna sequence from the hyperthermophilic crenarchaeon sulfolobus acidocaldarius is composed of two adjacent genes. the first functionally unassigned open reading frame (orf-1) comprises 450 base pairs. the second 1.1 kb large open reading frame encodes the putative signal recognition particle receptor alpha subunit (sr alpha). both genes are expressed under the heterotrophic growth conditions of the organism. the main transcript of orf-1 appears as a monocistronic rna in norther ...19968935656
bacteria breakfast. 19968959404
cloning and sequencing of a gene encoding 16s ribosomal rna from a novel hyperthermophilic archaebacterium nc12.a hyperthermophile nc12 was newly isolated from noboribetsu hot spring. to characterize this organism, a gene coding for 16s rrna was cloned and sequenced. the 16s rrna sequence from nc12 shows the highest similarity with those from pyrodictium occultum and desulfurococcus mobilis among the sequences in the database, indicating that nc12 belongs to a cluster of extreme thermophiles (crenarchaeota) in the archaeal domain. however, since the highest identity score was only 91.2%, it is suggested t ...19968973365
glutamate-1-semialdehyde aminotransferase from sulfolobus solfataricus.glutamate-1-semialdehyde aminotransferase (gsa-at) from the extremely thermophilic bacterium sulfolobus solfataricus has been purified to homogeneity and characterized. gsa-at is the last enzyme in the c5 pathway for the conversion of glutamate into the tetrapyrrole precursor delta-aminolaevulinate (ala) in plants, algae and several bacteria. the active form of gsa-at from s. solfataricus seems to be a homodimer with a molecular mass of 87 kda. the absorption spectrum of the purified aminotransf ...19968973563
hyperthermophile protein folding thermodynamics: differential scanning calorimetry and chemical denaturation of sac7d.recombinant sac7d protein from the thermoacidophile sulfolobus acidocaldarius is shown to be stable towards acid, thermal and chemical denaturation. the protein maintains a compact native fold between ph 0 and 10 in 0.3 m kcl and 25 degrees c as indicated by near and far uv circular dichroism spectra. thermal unfolding followed by differential scanning calorimetry (dsc) occurs as a reversible, two-state transition from ph 0 to 10, with a maximal tm of 90.7 degrees c between ph 5 and 9. at ph 0 t ...19968980686
bioenergetics of the archaebacterium sulfolobus.archaea are forming one of the three kingdoms defining the universal phylogenetic tree of living organisms. within itself this kingdom is heterogenous regarding the mechanisms for deriving energy from the environment for support of cellular functions. these comprise fermentative and chemolithotrophic pathways as well as light driven and respiratory energy conservation. due to their extreme growth conditions access to the molecular machineries of energy transduction in archaea can be experimental ...19968982385
kow: a novel motif linking a bacterial transcription factor with ribosomal proteins. 19968987397
gene cloning and expression of new trehalose-producing enzymes from the hyperthermophilic archaeum sulfolobus solfataricus km1.the genes encoding for trehalose-producing enzymes, a glycosyl-trehalose-producing enzyme (glycosyltransferase) and a gylcosyl-trehalose-hydrolyzing enzyme (alpha-amylase), from sulfolobus solfataricus km1 were cloned and expressed in e. coli. the nucleotide sequence of the glycosyltransferase gene and the alpha-amylase gene indicated proteins with lengths of 728 and 558 amino acids and molecular masses of 86-kda and 65-kda, respectively. regions highly conserved in the alpha-amylase family exis ...19968987868
immunochemical detection of adp-ribosylating enzymes in the archaeon sulfolobus solfataricus.polyclonal antibodies raised against eukaryotic mono-(adpribose)transferase and poly(adpribose)polymerase were used to test the presence of antigenic determinants in a crude extract of sulfolobus solfataricus, a thermophilic archaeon. samples from eukaryotic (bull testis) and bacterial (e. coli) sources were analysed for comparison. all tested antibodies reacted with the sulfolobal sample with a specificity comparable to that of the eukaryotic preparation, as revealed by elisa test, activity ass ...19968549833
cloning and overexpression in escherichia coli of the genes encoding nad-dependent alcohol dehydrogenase from two sulfolobus species.the gene adh encoding a nad-dependent alcohol dehydrogenase from the novel strain rc3 of sulfolobus sp. was cloned and sequenced. both the adh gene from sulfolobus sp. strain rc3 and the alcohol dehydrogenase gene from sulfolobus solfataricus (dsm 1617) were expressed at a high level in escherichia coli, and the recombinant enzymes were purified, characterized, and compared. only a few amino acid replacements were responsible for the different kinetic and physicochemical features investigated.19968550434
isolation, characterization and crystallization of an iron-superoxide dismutase from the crenarchaeon sulfolobus acidocaldarius.an iron containing superoxide dismutase from the cytosol of the thermoacidophilic crenarchaeon sulfolobus acidocaldarius (dsm 639) has been purified to electrophoretic homogeneity. it comprises at least 11% of the cytosolic protein. the isolated protein consists of two identical subunits with an apparent molecular mass of 22.4 kda. it contains one iron atom per dimer. the protein shows the typical epr spectrum of a s = 3/2, rhombic high-spin iron center. it is extremely resistant against thermal ...19968674972
isolation and characterization of flagella and flagellin proteins from the thermoacidophilic archaea thermoplasma volcanium and sulfolobus shibatae.isolated flagellar filaments of sulfolobus shibatae were 15 nm in diameter, and they were composed of two major flagellins which have m(r)s of 31,000 and 33,000 and which stained positively for glycoprotein. the flagellar filaments of thermoplasma volcanium were 12 nm in diameter and were composed of one major flagellin which has an m(r) of 41,000 and which also stained positively for glycoprotein. n-terminal amino acid sequencing indicated that 18 of the n-terminal 20 amino acid positions of th ...19968550530
dtmp biosynthesis in archaea.the biosynthesis of dtmp has been studied in cell extracts of two different members of the domain archaea, methanosarcina thermophila and sulfolobus solfataricus. in m. thermophila, the dtmp was formed from dump and [methylene-2h2]-5,10-methylenetetrahydrosarcinapterin generated in situ from added [methylene-2h2] formaldehyde and the tetrahydrosarcinapterin present in the cell extract. in s. solfataricus, the 5,10-methyl-enetetrahydro derivative of a synthetic fragment of sulfopterin, the modifi ...19968550533
complete nucleotide sequence of the sulfolobus islandicus multicopy plasmid prn1.the complete sequence of the 5350-bp plasmid prn1 from the crenarchaeote sulfolobus islandicus has been determined. this plasmid is the first to be sequenced from this group of thermoacidophilic archaebacteria (archaea) and its high copy number and wide host range make it a good candidate for a cloning vector. prn1 contains several open reading frames, including one that spans over half the plasmid and has significant similarity to the helicase domain of viral primase proteins. directly upstream ...19968700967
activation of sulfolobus solfataricus alcohol dehydrogenase by modification of cysteine residue 38 with iodoacetic acid.reaction of thermostable nad(+)-dependent alcohol dehydrogenase from sulfolobus solfataricus with iodoacetate at ph 9.0 and 37 degrees c significantly increases the oxidation rate of aliphatic and aromatic alcohols and decreases the reduction rate of aromatic aldehydes. the archaeal adh is chemically modified and activated in a michaelis-menten-type reaction, where one molecule of the reagent binds per active site. nad+ in micromolar concentration protects the enzyme against the inhibitor in an ...19968555238
comparative analysis of ribonuclease p rna structure in archaea.although the structure of the catalytic rna component of ribonuclease p has been well characterized in bacteria, it has been little studied in other organisms, such as the archaea. we have determined the sequences encoding rnase p rna in eight euryarchaeal species: halococcus morrhuae, natronobacterium gregoryi, halobacterium cutirubrum, halobacteriurn trapanicum, methanobacterium thermoautotrophicum strains deltah and marburg, methanothermus fervidus and thermococcus celer strain al-1. on the b ...19968614627
cloning of a potential cytochrome p450 from the archaeon sulfolobus solfataricus.abstract a gene, cyp119, for a potential cytochrome p450 has been isolated and sequenced from the extreme acidothermophilic archaeon sulfolobus solfataricus. the gene predicts a polypeptide of 368 amino acids containing the consensus heme-binding sequence phe-gly-xaa-gly-xaa-his-xaa-cys-xaa-gly- xaa3-ala-arg-xaa-glu. it most closely resembles the cytochrome p450s found in the bacterium bacillus subtilis, with which it shares 129 identical amino acid residues (35%). this first sequence of a poten ...19968617361
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