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Biologically produced alcohols are of great current interest for renewable solvents and liquid transportation fuels. While bioethanol is now produced on a massive scale, butanol has superior fuel characteristics and an additional value as a solvent and chemical feedstock. Butanol production has been demonstrated at ambient temperatures in metabolically-engineered mesophilic organisms, but the ability to engineer a microbe for in vivo high-temperature production of commodity chemicals has several distinct advantages. These include reduced contamination risk, facilitated removal of volatile products, and a wide temperature range to modulate and balance both the engineered pathway and the host׳s metabolism. We describe a synthetic metabolic pathway assembled from genes obtained from three different sources for conversion of acetyl-CoA to 1-butanol, and 1-butanol generation from glucose was demonstrated near 70 °C in a microorganism that grows optimally near 100 °C. The module could also be used in thermophiles capable of degrading plant biomass. 相似文献
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Jocelyne DiRuggiero James R. Brown Allison P. Bogert Frank T. Robb 《Journal of molecular evolution》1999,49(4):474-484
DNA repair in the Archaea is relevant to the consideration of genome maintenance and replication fidelity in the last universal
common ancestor (LUCA) from two perspectives. First, these prokaryotes embody a mix of bacterial and eukaryal molecular features.
Second, DNA repair proteins would have been essential in LUCA to maintain genome integrity, regardless of the environmental
temperature. Yet we know very little of the basic molecular mechanisms of DNA damage and repair in the Archaea in general.
Many studies on DNA repair in archaea have been conducted with hyperthermophiles because of the additional stress imposed
on their macromolecules by high temperatures. In addition, of the six complete archaeal genome sequences published so far,
five are thermophilic archaea. We have recently shown that the hyperthermophile Pyrococcus furiosus has an extraordinarily high capacity for repair of radiation-induced double-strand breaks and we have identified and sequenced
several genes involved in DNA repair in P. furiosus. At the sequence level, only a few genes share homology with known bacterial repair genes. For instance, our phylogenetic
analysis indicates that archaeal recombinases occur in two paralogous gene families, one of which is very deeply branched,
and both recombinases are more closely related to the eukaryotic RAD51 and Dmc1 gene families than to the Escherichia coli recA gene. We have also identified a gene encoding a repair endo/exonuclease in the genomes of several Archaea. The archaeal sequences
are highly homologous to those of the eukaryotic Rad2 family and they cluster with genes of the FEN-1 subfamily, which are
known to be involved in DNA replication and repair in eukaryotes. We argue that there is a commonality of mechanisms and protein
sequences, shared between prokaryotes and eukaryotes for several modes of DNA repair, reflecting diversification from a minimal
set of genes thought to represent the genome of the LUCA. 相似文献
75.
Takaaki Sato Masahiro Fujihashi Yukika Miyamoto Keiko Kuwata Eriko Kusaka Haruo Fujita Kunio Miki Haruyuki Atomi 《The Journal of biological chemistry》2013,288(29):20856-20867
Here we performed structural and biochemical analyses on the TK2285 gene product, an uncharacterized protein annotated as a member of the ribokinase family, from the hyperthermophilic archaeon Thermococcus kodakarensis. The three-dimensional structure of the TK2285 protein resembled those of previously characterized members of the ribokinase family including ribokinase, adenosine kinase, and phosphofructokinase. Conserved residues characteristic of this protein family were located in a cleft of the TK2285 protein as in other members whose structures have been determined. We thus examined the kinase activity of the TK2285 protein toward various sugars recognized by well characterized ribokinase family members. Although activity with sugar phosphates and nucleosides was not detected, kinase activity was observed toward d-allose, d-lyxose, d-tagatose, d-talose, d-xylose, and d-xylulose. Kinetic analyses with the six sugar substrates revealed high Km values, suggesting that they were not the true physiological substrates. By examining activity toward amino sugars, sugar alcohols, and disaccharides, we found that the TK2285 protein exhibited prominent kinase activity toward myo-inositol. Kinetic analyses with myo-inositol revealed a greater kcat and much lower Km value than those obtained with the monosaccharides, resulting in over a 2,000-fold increase in kcat/Km values. TK2285 homologs are distributed among members of Thermococcales, and in most species, the gene is positioned close to a myo-inositol monophosphate synthase gene. Our results suggest the presence of a novel subfamily of the ribokinase family whose members are present in Archaea and recognize myo-inositol as a substrate. 相似文献
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Abstract The phosphoglycerate kinase ( pgk ), triosephosphate isomerase ( tpi ), and enolase ( eno ) genes from Thermotoga neapolitana have been cloned and expressed in Escherichia coli . In high copy number, the pgk gene complemented an E. coli pgk − strain. In T. neapolitana , the pgk and tpi genes appear to be fused and eno is near those genes. Like T. maritima , T. neapolitana produces phosphoglycerate kinase as both an individual enzyme and a fusion protein with triosephosphate isomerase, and triosephosphate isomerase activity is not found without associated phosphoglycerate kinase activity. Unlike T. maritima , which forms only a 70-kDa fusion protein, T. neapolitana expresses both 73-kDa and 81-kDa isozymes of this fusion protein. These isozymes are present in both T. neapolitana cells and in E. coli cells expressing T. neapolitana genes. 相似文献
78.
Kecha M Benallaoua S Touzel JP Bonaly R Duchiron F 《Extremophiles : life under extreme conditions》2007,11(1):65-73
A hyperthermophilic anaerobic archeon, strain HT3, was isolated from hydrothermal hot spring in Northeast Algeria. The strain
is a regular coccus, highly motile, obligatory anaerobic, heterotrophic. It utilizes proteinaceous complex media (peptone,
tryptone or yeast extract). Sulfur is reduced to Hydrogen sulfide and enhances growth. It shares with other Pyrococcus species the heterotrophic mode of nutrition, the hyperthermophily, the ability to utilize amino acids as sole carbon and nitrogen
sources and the ether lipid composition. The optimal growth occurs at 80–85°C, pH 7.5 and 1.5% NaCl. The G + C content was
43 mol%. Considering its morphology, physiological properties, nutritional features and phylogenetic analyses based on 16S
rRNA gene sequencing, this strain is described as a new terrestrial isolate pertaining to the genus Pyrococcus. 相似文献
79.
Angels Tapias Christophe Leplat Fabrice Confalonieri 《Extremophiles : life under extreme conditions》2009,13(2):333-343
The recently discovered hyperthermophilic and radioresistant archaeon Thermococcus gammatolerans is of great interest to compare and contrast the impact of its physiology on radioresistance and its ability to repair damaged
chromosomes after exposure to gamma irradiation with radioresistant bacteria. We showed that, in contrast to other organisms,
cell survival was not modified by the cellular growth phase under optimal growth conditions but nutrient-limited conditions
did affect the T. gammatolerans radioresistance. We determined the first kinetics of damaged DNA recovery in an archaeon after exposure to massive doses
of gamma irradiation and compared the efficiency of chromosomal DNA repair according to the cellular growth phase, nutrient
availability and culture conditions. Chromosomal DNA repair kinetics showed that stationary phase cells reconstitute disrupted
chromosomes more rapidly than exponential phase cells. Our data also revealed that this radioresistant archaeon was proficient
to reconstitute shattered chromosomes either slowly or rapidly without any loss of viability. These results suggest that rapid
DNA repair is not required for the extreme radioresistance of T. gammatolerans.
Angels Tapias and Christophe Leplat contributed equally to this work. 相似文献
80.
Elemental sulfur reduction by the hyperthermophilic bacterium Thermotoga neapolitana provides an alternative to hydrogen evolution during fermentation. Electrons are transferred from reduced cofactors (ferredoxin
and NADH) to sulfur by a series of unknown steps. One enzyme that may be involved is an NADH:methyl viologen oxidoreductase
(NMOR), an activity that in other fermenting organisms is associated with NADH:ferredoxin oxidoreductase. We found that 83%
of NMOR activity was contained in the pellet fraction of cell extracts subjected to ultracentrifugation. This pellet fraction,
presumably containing cell membranes, was required for electron transfer to NAD+ from ferredoxin-dependent pyruvate oxidation. However, the NMOR activity in this fraction used neither Thermotoga nor clostridial ferredoxins as substrates. NMOR activity was also detected in aerobically prepared vesicles. By comparison
with ATPase activities, NMOR was found primarily on the cytoplasmic face of these vesicles. During these studies, an extracytoplasmic
hydrogenase activity was discovered. In contrast to the soluble hydrogenase, this hydrogenase activity was completely inhibited
when intact cells were treated with cupric chloride and was present on the extracytoplasmic face of vescides. In contrast
to a soluble hydrogenase reported in Thermotoga maritima, this activity was air-stable and was inhibited by low concentrations of nitrite.
Received: 28 May 1998 / Accepted: 23 June 1998 相似文献