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  • 1
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The enzyme activities of the pentose phosphate pathway in the ethanologenic, Gram-negative bacterium Zymomonas mobilis were studied in order to construct a xylose catabolic pathway. In cell-free extracts of wild-type Z. mobilis CP4, activities of the enzymes transketolase (TKT) [2 munits (U)/mg], phosphoribose epimerase (640 mU/mg), phosphoribose isomerase (1600 mU/mg) and 6-phosphogluconate dehydrogenase (2 mU/mg) were determined. However, no transaldolase activity could be detected. Recombinant strains of Z. mobilis were constructed that carried the xylAB genes of the xylose catabolic pathway from Klebsiella pneumoniae. Expression of xylose isomerase (XI, 150 mU/mg) and xylulokinase (XK) (1300 mU/mg) were found in recombinant strains but no growth on pentose as sole carbon source occurred. The xyl-recombinant cells were moreover growth-inhibited in the presence of xylose and were found to accumulate xylitol phosphate due to the subsequent action of a novel enzyme, an NADPH-dependent aldose reductase, and a side reaction of XK on xylitol. From the xylAB recombinant strains, mutants were isolated that were less inhibited and formed less xylitol phosphate when grown in the presence of xylose. The tkt gene of E. coli was cloned on the xylAB plasmid and introduced into Z. mobilis strains. This led to higher TKT activities (150 mU/mg) and, in cooperation with the enzymes XI and XK, mediated a conversion of small amounts of xylose to CO2 and ethanol. However, no growth on xylose as sole carbon source was detected, instead sedoheptulose 7-P accumulated intracellularly.
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  • 2
    ISSN: 1432-072X
    Keywords: d-Fructose ; New pathway ; l-Sorbose ; E. coli ; K. pneumoniae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Starting with a fruK (formerly fpk) mutant of Escherichia coli K12 lacking d-fructose-1-phosphate kinase (E.C. 2.7.1.3.), fructose positive derivatives were isolated after introduction of the cloned gene sorE from Klebsiella pneumoniae coding for an l-sorbose-1-phosphate reductase. The new pathway was shwon to proceed from d-fructose via d-fructose-1-phosphate and d-mannitol-1-phosphate to d-fructose 6-phosphate. It involves a transport system and enzymes encoded in the fru and the mtl operons from E. coli K12 as well as in the sor operon from K. pneumoniae respectively.
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  • 3
    ISSN: 1617-4623
    Keywords: Klebsiella pneumoniae ; Xylose isomerase ; Xylulokinase ; xy1 gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The genes xy1A and xy1B were cloned together with their promoter region from the chromosome of Klehsiella pneumoniae var. aerogenes 1033 and the DNA sequence (3225 bp) was determined. The gene xy1A encodes the enzyme xylose isomerase (XI or XylA) consisting of 440 amino acids (calculated Mr of 49 793). The gene xy1B encodes the enzyme xylulokinase (XK or Xy1B) with a calculated M, of 51 783 (483 amino acids). The two genes successfully complemented xy1 mutants of Escherichia coli K12, but no gene dosage effect was detected. E. coli wild-type cells which harbored plasmids with the intact xylA Kp 5′ upstream region in high copy number (but lacking an active xy1B gene on the plasmids) were phenotypically xylose-negative and xylose isomerase and xylulokinase activities were drastically diminished. Deletion of 5′ upstream regions of xy1A on these plasmids and their substitution by a lac promoter resulted in a xylose-positive phenotype. This also resulted in overproduction of plasmid-encoded xylose isomerase and xylulokinase activities in recombinant E. coli cells.
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  • 4
    ISSN: 1617-4623
    Keywords: Pyruvate decarboxylase gene (pdc) ; Expression vectors ; Promoter/terminator ; Chloramphenicol acetyltransferase ; Zymomonas mobilis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A set of vectors was constructed for the cloning and expression of heterologous genes in the Gramnegative bacterium Zymomonas mobilis under the control of the pdc promoter of Z. mobilis. The vectors pPTZ1, pPTZ3, and pPTZ4 are based on the cryptic Z. mobilis plasmid pZM02 and on parts of the Escherichia coli plasmids pKK223-3 and pBR322 together with the multiple cloning site of phage Ml3mp18. DNA fragments can be readily inserted immediately downstream from the pdc promoter at unique restriction sites for KpnI, XbaI and PstI in pPTZl and additionally for SmaI and BamHI in pPTZ3. In pPTZ4, the 5′ terminal codons of pdc were deleted allowing the formation of gene fusions. Expression of a promoterless chloramphenicol acetyltransferase gene (cat) controlled by the pdc gene promoter resulted in enzyme activities of up to 5.5 U/mg total cell protein in Z. mobilis cells.
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  • 5
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] TABLE 1 Vibrational parameters for silicon clusters Experimental Calculated Absolute frequency frequency deviation Symmetry Calculated Cluster (cm"1) (cm"1)* (cm"1) assignment intensityt Si4 345 337 8 Ag 1.0 440 B3g 0.5 470 463 7 A* 5.0 Si6J 252 209 43 B2g 1.0 300 298 2 A* 2.6 ...
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  • 6
    Electronic Resource
    Electronic Resource
    [s.l.] : Nature Publishing Group
    Nature 360 (1992), S. 600-603 
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Maternally derived Raf protein is required during embry-ogenesis for the response to activation of the torso receptor tyrosine kinase13. Strong loss-of-function raf mutations block cell proliferation11'12, and as anticipated, our attempts to examine the consequences of complete loss of raf function ...
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  • 7
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract The enzyme glucose-fructose oxidoreductase (GFOR) from the Gram-negative ethanologenic bacterium Zymomonas mobilis was purified to homogeneity and was shown to be a tetrameric protein with a subunit size of Mr 42 500. Using immunogold-labelling in combination with electron microscopy, ultrathin sections of Z. mobilis wild type cells showed that the enzyme GFOR is located in the periplasm off the bacterial cells. Z. mobilis strains which carried the cloned gfo gene on plasmid pSUP104, had 5–6-fold increased GFOR enzyme activities. Moreover, these cells accumulated large amounts of a presumable unprocessed pre-GFOR protein (Mr 48 000).
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  • 8
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract Glucose-fructose oxidoreductase (GFOR) is a periplasmic enzyme of the ethanologenic, Gram-negative bacterium Zymomonas mobilis. It contains tightly bound NADP+ as cofactor. In Z. mobilis GFOR-recombinant strains, a precursor form of GFOR was accumulated. To assay the preGFOR for its NADP(H) content and enzymatic activity, it was purified from an overproducing strain. Using SDS-PAGE, the precursor subunit size was determined to approximately 45 kDa (compared with a 40 kDa subunit size for the mature GFOR subunit). The N-terminal amino acid sequence of the precursor was determined. The N-terminal residues of the GFOR matched with the signal sequence from the DNA sequence of the gene gfo. The precursor form of GFOR was enzymatically active and contained the cofactor NADP(H).
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  • 9
    ISSN: 1432-1459
    Keywords: Inflammatory polyneuropathy ; Demyelination ; Cyclosporin A ; Plasma exchange
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary A patient with chronic relapsing inflammatory demyelinating polyneuropathy was successfully treated with plasma exchanges and cyclosporin A (CsA). Dynamometric measurements of hand force during the time of CsA treatment showed a highly significant correlation between hand force and CsA blood levels. The largest influence of CsA on hand force occurred 11–13 days after CsA uptake.
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  • 10
    ISSN: 1432-1440
    Keywords: Activated charcoal ; Sorbitol ; Paracetamol ; Diphenhydramine ; Codeine ; Adsorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Due to its paramount adsorption capacity, activated charcoal is supposed to be the remedy of choice for binding a variety of drugs in the gastrointestinal tract. Hence it is surprising — at least according to the advice of German textbooks — that activated charcoal is only recommended for administration after time-consuming treatments like induced emesis and gastric lavage. Particularly with infants at home, a ready-for-use suspension of activated charcoal would allow the early management of acute poisoning. In such cases, inactivation of the poison by adsorption could be particularly helpful, since the period after ingestion is usually short. The charcoal-sorbitol-suspension (30 g activated charcoal in 150 ml of 70% sorbitol) is a creamy preparation which is easy to drink, because density and viscosity prevent sedimentation. The prescription-free drugs can be dispensed by each pharmacist. The present study was undertaken to investigate the influence of sorbitol on the adsorption capacity of activated charcoal. To this end, adsorption isotherms were established in vitro and compared with results in volunteers to whom NAPAP, diphenhydramine or codeine was administered separately. These drugs are gaining increasing importance in medicinal toxicology since they are constituents of various analgesics and cold remedies. To determine absorption, the cumulative urinary excretion was estimated of the parent drugs and their main metabolites. Without charcoal the volunteers were dosed with 500 mg NAPAP, 50 mg diphenhydramine-HCl, and 50 mg codeine phosphate, respectively. When the charcoal-sorbitol-suspension was swallowed two min after ingestion of the tablett-slurry, the five-fold dose of each drug was administered. The charcoal-sorbitol-suspension significantly (p〈0.01) diminished systemic absorption of codeine to 12%, of diphenhydramine to 28% and of NAPAP to 44% (means, n=5). These results demonstrate the effectiveness of the charcoal-sorbitol-suspension to inhibit drug absorption. In vitro, sorbitol at neutral and acidic pH reduced the maximal adsorbance capacity of charcoal mainly for codeine (by 72%), less for diphenhydramine (19%) and least for NA-PAP (14%). Hence it is obvious that in-vitro experiments hardly predict the efficacy in vivo. This discrepancy suggests that part of the drugs had been already absorbed before binding to charcoal. This assumption is underlined by one experiment when diphenhydramine and charcoal-sorbitol were taken together. In this case less than 2% of the drug was absorbed. The charcoal-sorbitol-suspension was generally well tolerated besides marked flatulence. Black stools appeared within 1–3 h, the laxative action persisted for about 8 hours. It is recommended to administer activated charcoal to acutely poisoned patients as soon as possible (children 1 g/kg). The only exceptions are obtunded patients or ingestion of caustics. This first aid measure should start immediately at home or at work (e.g. after advice by telephone), in the ambulance car, and in busy (lack of staff !) intensive care units. When charcoal is rather ineffective (alcohol, iron or lithium salts) or in case of very large quantities of the ingested poison, emesis and gastric lavage can be performed even after charcoal administration.
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