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1.
Growing cultures of Acetobacter melanogenus ATCC 9937 concerted D-glucose to 2,5-diketo-D-gluconic acid with D-gluconic acid and 5-keto-D-gluconic acid as intermediates. The 2,5-diketo-D-gluconic acid was isolated from the fermented medium by treatment with an anion exchange resin. 相似文献
2.
Preparation of Cells Permeable to Macromolecules by Treatment with Toluene: Studies of Transfer Ribonucleic Acid Nucleotidyltransferase 总被引:2,自引:1,他引:1
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Murray P. Deutscher 《Journal of bacteriology》1974,118(2):633-639
Transfer ribonucleic acid (tRNA) nucleotidyltransferase was studied after making cells permeable to macromolecules by treatment with toluene. The conditions of toluene treatment necessary for obtaining maximal activity were defined. Toluene treatment was most efficient when carried out for 5 min at 37 C at pH 9.0 on log-phase cells. No activity could be detected if cells were treated at 0 C, or in the presence of MgCl2, or if the cells were in the stationary phase of growth. However, inclusion of lysozyme and ethylenediaminetetraacetic acid during the toluene treatment did render stationary phase cells permeable. The properties of tRNA nucleotidyltransferase from toluene-treated cells were essentially identical to those of purified enzyme with regard to pH optimum, specificity for nucleoside triphosphates and tRNA, and apparent Km values for substrates. In addition to tRNA nucleotidyltransferase, a variety of other enzymes which incorporate adenosine 5′-triphosphate into acid-precipitable material could also be detected in toluene-treated cells. Centrifugation of cells treated with toluene revealed that tRNA nucleotidyltransferase leaked out of cells, whereas other activities remained associated with the cell pellets. Chromatography of the material extracted from toluene-treated cells on Sephadex G-100 indicated that toluene treatment selectively extracts lower molecular weight proteins. The usefulness of such a procedure as an initial step in purification of such enzymes, and its application to tRNA nucleotidyltransferase, is discussed. 相似文献
3.
Degradation of Toluene and Trichloroethylene by Burkholderia cepacia G4 in Growth-Limited Fed-Batch Culture 总被引:1,自引:0,他引:1
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Burkholderia (Pseudomonas) cepacia G4 was cultivated in a fed-batch bioreactor on either toluene or toluene plus trichloroethylene (TCE). The culture was allowed to reach a constant cell density under conditions in which the amount of toluene supplied equals the maintenance energy demand of the culture. Compared with toluene only, the presence of TCE at a toluene/TCE ratio of 2.3 caused a fourfold increase in the specific maintenance requirement for toluene from 22 to 94 nmol mg of cells (dry weight)(sup-1) h(sup-1). During a period of 3 weeks, approximately 65% of the incoming TCE was stably converted to unidentified products from which all three chlorine atoms were liberated. When toluene was subsequently omitted from the culture feed while TCE addition continued, mutants which were no longer able to grow on toluene or to degrade TCE appeared. These mutants were also unable to grow on phenol or m- or o-cresol but were still able to grow on catechol and benzoate. Plasmid analysis showed that the mutants had lost the plasmid involved in toluene monooxygenase formation (pTOM). Thus, although strain G4 is much less sensitive to TCE toxicity than methanotrophs, deleterious effects may still occur, namely, an increased maintenance energy demand in the presence of toluene and plasmid loss when no toluene is added. 相似文献
4.
Summary The effect of permeabilization of G. suboxydans cells with toluene on the oxidation of D-sorbitol to L-sorbose was investigated. Treatment of the cells with 10% toluene resulted in a three fold increase in the specific sorbitol dehydrogenase activity and a two fold increase in the efficiency of D-sorbitol conversion to L-sorbose of the free cell suspension. When the permeabilized cells were immobilized in calcium alginate, the operational stability during air-lift reactor operation was also found to increase with up to three times longer half-life(44 days) of catalytic activity compared with immobilized intact cells. 相似文献
5.
R H Hilderman P J Goldblatt M P Deutscher 《The Journal of biological chemistry》1975,250(12):4796-4801
Isolated individual liver cells were made permeable to charged molecules and macromolecules by treatment with toluene, and the properties of such cells were examined in detail. The optimal conditions of toluene treatment, as determined by assay of aminoacyl-tRNA synthetase activities, was 7 to 9% toluene for 2 min at 0 degrees. Conditions are also described for maintaining the treated cells for periods up to 1 hour. Toluene treatment was found to be as efficient as various disruptive methods for making internal enzymes accessible to exogenous substrates, and this was true for enzymes in the cytosol, membrane, or organelle fractions. Electron microscopic and biochemical examination of toluen-treated cells indicated that they were relatively intact and lost only small amounts of cellular constituents to the suspension medium. The data in this paper suggest that toluene treatment of individual cells might prove useful for studies of macromolecular synthesis in liver; 相似文献
6.
S Makover G B Ramsey F M Vane C G Witt R B Wright 《Biotechnology and bioengineering》1975,17(10):1485-1514
L-Sorbose is oxidized to 2-keto-L-gulonic acid (KGA) via the following sequence of reactions which we call the "sorbosone pathway": L-sorbose in equilibrium L-sorbosone leads to KGA. The first step is reversible and is mediated by enzymes found in a soluble fraction obtained from Pseudomonas putida ATCC 21812. Although no cofactor requirements were found for the forward reaction, the reverse reaction clearly required NADH. Enzymes for this NADH-dependent synthesis of L-sorbose could be differentiated on the basis of molecular weights. The second step in the sorbosone pathway is catalyzed by a particulate enzyme found in extracts from P. putida and Gluconobacter melanogenus IFO 3293. The rate limiting reaction in the sorbosone pathway is the synthesis of L-sorbosone. In addition to P. putida, Klebsiella pneumoniae (ATCC 27858) and Serratia marcescens (ATCC 27857) also contain the enzymes which catalyze the reactions of the sorbosone pathway. Two of the bacteria studied, P. putida and G. melanogenus, also contain an enzyme involved in the further metabolism of KGA to L-idonic acid. This enzyme, referred to as KGA-reductase, is found in the soluble fraction of cell-free extracts and is dependent on NADH or NADPH. 相似文献
7.
The regulation of aspartokinase and homoserine dehydrogenase has been studied in three Acetobacter and two Gluconobacter species. Both enzymes were regulated by feedback inhibition. Aspartokinase was inhibited by L-threonine and concertedly inhibited by L-threonine plus L-lysine. The homoserine dehydrogenase was NADP-specific and was inhibited by L-threonine. Separation of the two enzymes by ammonium sulphate fractionation was possible in Acetobacter peroxydans, A. rancens and Gluconobacter melanogenus but not in A. liquefaciens or G. oxydans. 相似文献
8.
Toluene-induced cells of Pseudomonas putida NCIMB 11767 lost their ability to oxidise toluene within 300 h under conditions of carbon/energy or nitrogen deprivation at 30°C, while incubation at 4°C improved the stability of this activity. Provision of inducing substrates (toluene or phenol) to nitrogen-deprived cells at 30°C also enhanced the stability of toluene oxidation, whereas provision of a non-inducing carbon/energy source (ethanol) led to a total loss of toluene oxidation within 160 h. Disappearance of toluene-induced proteins, at different rates accompanied the loss of toluene oxidation in carbon-deprived cells. The data suggest that degradation of one or more of the major proteins of toluene metabolism determines the stability of toluene oxidation in carbon-deprived cells. Around 40% of the whole-cell toluene oxidation rate was recoverable after cryopreservation (–20°C under glycerol) of toluene-induced cells but most of this recovered activity (86%) was associated with dead cells. These observations may have important implications for the application of these toluene-induced cells as in situ bioremediation catalysts. 相似文献
9.
Permanent loss of chromosome initiation in toluene-treated Bacillus subtilis cells. 总被引:2,自引:1,他引:1
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Initiation of deoxyribonucleic acid replication is absent in Bacillus subtilis cells made permeable by toluene. The absence of initiation may be (i) a temporary removal of toluene, or (ii) irreversibly lost due to damage by toluene treatment to a cellular structure or a process required for chromosome initiation. Washed cells, previously treated with toluene and subsequently washed free of detectable amounts of toluene, have the same characteristics of toluene-treated cells in which toluene is not removed. The continued absence of initiation in the washed cells indicates a permanent loss of initiation in cells treated with toluene. Protein synthesis is also inhibited irreversibly by toluene treatment, indicating damage to translation as a possible mechanism for loss of initiation of toluene-treated cells. 相似文献
10.
Gluconobacter melanogenus IFO 3293 cells capable of converting L-sorbose to L-sorbosone were immobilized in polyacrylamide gel. The preferred polymer composition for high activity and stability was determined to contain a total monomer concentration of 7.2% and 16.6% crosslinking agent. No significant differences in optimal conditions for conversion, e.g., pH and temperature, were found in comparison with free cell suspensions. However, in the absence of L-sorbose, the thermal stability of immobilized cells was lower. After the initial loss, the conversion activity of immobilized cells increased, possibly due to lysis, and this increase was related to the polymerization conditions and the incubation temperature for the L-sorbose conversion. The enzymatic activity and stability of the immobilized cells also depended on the physical form of the gel and the aeration levels. Addition of electron acceptors or addition of L-sorbosone to the medium reduced, while addition of neomycin, ampicillin, chloramphenicol, and tetracycline increased the stability of the enzymatic activity. 相似文献
11.
Physiological stress in batch cultures of Pseudomonas putida 54G during toluene degradation 总被引:1,自引:0,他引:1
R G Mirpuri W L Jones G A McFeters H F Ridgway 《Journal of industrial microbiology & biotechnology》1997,18(6):406-413
Physiological stress associated with toluene exposure in batch cultures of Pseudomonas putida 54G was investigated. P. putida 54G cells were grown using a continuous vapor phase feed stream containing 150 ppmv or 750 ppmv toluene as the sole carbon
and energy source. Cells were enumerated on non-selective (R2A agar plates) and a selective minimal medium incubated in the
presence of vapor phase toluene (HCMM2). Differential recovery on the two media was used to evaluate bacterial stress, culturability
and loss of toluene-degrading capability. A majority of the bacteria were reversibly stressed and could resume active colony
formation on selective medium after passage on non-selective medium. A small fraction of the bacterial cells suffered an
irreversible loss of toluene degradation capability and were designated as Tol− variants. Numbers of stressed organisms increased with duration of toluene exposure and toluene concentration and coincided
with accumulation of metabolic intermediates from incomplete toluene degradation. Respiring cell numbers in the batch cultures
decreased as injury increased, indicating a possible relationship between respiring and injured cells. Rate expressions for
injury, for formation of Tol− variants and for growth of Tol− variants were determined by calibrating a theoretical model to the results obtained. These rate expressions can be used
to calibrate bioreactor models, and provide a basis for better design and control of bioremediation systems.
Received 01 July 1996/ Accepted in revised form 25 March 1997 相似文献
12.
Alteration of the Cloudman melanoma cell cycle by prostaglandins E1 and E2 determined by using a 5-bromo-2'-deoxyuridine method of DNA analysis 总被引:1,自引:0,他引:1
Z A Abdel-Malek V B Swope L S Trinkle E N Ferroni R E Boissy J J Nordlund 《Journal of cellular physiology》1988,136(2):247-256
Prostaglandins (PGs) E1 and E2 stimulate tyrosinase activity and suppress the proliferation of Cloudman S91 melanoma cells by altering their progression through the cell cycle. Prostaglandin E1 and PGE2 have prolonged or residual effects on melanoma cells. Cells treated for 5 or 24 hours with 10 micrograms/ml PGE1 or cells treated for 8 or 24 hours with 10 micrograms/ml PGE2 demonstrated decreased proliferation and increased tyrosinase activity for 48 hours after removal of the PGs. The effects of PGs on the cell cycle were investigated by determining total DNA content in cells stained with propidium iodide (PI) and analyzed by a fluorescence activated cell sorter (FACS). Prostaglandin E1 blocked cells in G2 phase after 5 hours of treatment, corresponding to when inhibition of proliferation was first evident. Similarly, after 9 hours of treatment with PGE2, more cells were in late S, early G2 phase and less in G1 than their control counterparts. Also, melanoma cells were pulse-labeled with 5-bromo-2'-deoxyuridine (BrdUrd) prior to or at the end of PG treatment and then stained with a fluoresceinated monoclonal antibody to BrdUrd, and with PI. This allows one to observe how BrdUrd-labeled S-phase cells cycle with time. Both PGE1 and PGE2 inhibit proliferation by blocking cells in G2 phase of the cell cycle. The PG-induced block in G2 may be required by melanoma cells to synthesize mRNA and proteins that are essential for stimulation of tyrosinase activity. Ultrastructurally, only a subpopulation of the cells treated with PGE1 or PGE2 contained more mature melanosomes than control cells. 相似文献
13.
The effect of toluene on the structure and permeability of the outer and cytoplasmic membranes of Escherichia coli. 总被引:8,自引:0,他引:8
The effect of toluene on Escherichia coli has been examined. In the presence of Mg2+, toluene removes very little protein, phospholipid, or lipopolysacharide from E. coli. In the absence of Mg2+, or in the presence of EDTA, toluene removes considerably more cell material, including several specific cytoplasmic proteins such as malate dehydrogenase (EC 1.1.1.37). In contrast, glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and glutamate dehydrogenase (EC 1.4.1.4) are not released at all under the same conditions. Cells treated with toluene in the presence of Mg2+ remain relatively impermeable to pyridne nucleotides, while cells treated with toluene in the presence of EDTA become permeable to these compounds. Freeze-fracture electron microscopy shows that toluene causes considerable damage to the cytoplasmic membrane, while the outer membrane remains relatively intact. These results indicate that the permeability characteristics of toluene-treated cells depend at least partly on the state of the outer membrane after the toluene treatment. 相似文献
14.
Kanter M 《Neurochemical research》2008,33(11):2241-2249
The goal of this study was designed to evaluate the possible protective effects of Nigella sativa (NS) on the neuronal injury in the frontal cortex and brain stem after chronic toluene exposure in rats. The rats were randomly
alotted into one of three experimental groups: A (control), B (toluene treated) and C (toluene treated with NS); each group
contain 10 animals. Control group received 1 ml serum physiologic and toluene treatment was performed by inhalation of 3,000 ppm
toluene, in a 8 h/day and 6 day/week order for 12 weeks. The rats in NS treated group was given NS (in a dose of 400 mg/kg
body weight) once a day orally by using intra gastric intubation for 12 weeks starting just after toluene exposure. Tissue
samples were obtained for histopathological investigation. To date, no histopathological changes of neurodegeneration in the
frontal cortex and brain stem after chronic toluene exposure in rats by NS treatment have been reported. In this study, chronic
toluene exposure caused severe degenerative changes, shrunken cytoplasma, severely dilated cisternae of endoplasmic reticulum,
markedly swollen mitochondria with degenerated cristae and nuclear membrane breakdown with chromatin disorganization in neurons
of the frontal cortex and brain stem. The nerve cells showing the pathologic changes were almost absent in the NS-treated
rats. We conclude that NS therapy causes morphologic improvement on neurodegeneration in frontal cortex and brain stem after
chronic toluene exposure in rats. We believe that further preclinical research into the utility of NS may indicate its usefulness
as a potential treatment on neurodegeneration after chronic toluene exposure in rats. 相似文献
15.
Biochemical Localization of Alkaline Phosphatase in the Cell Wall of a Marine Pseudomonad 总被引:8,自引:5,他引:3
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The various layers of the cell envelope of marine pseudomonad B-16 (ATCC 19855) have been separated from the cells and assayed directly for alkaline phosphatase activity under conditions established previously to be optimum for maintenance of the activity of the enzyme. Under conditions known to lead to the release of the contents of the periplasmic space from the cells, over 90% of the alkaline phosphatase was released into the medium. Neither the loosely bound outer layer nor the outer double-track layer (cell wall membrane) showed significant activity. A small amount of the alkaline phosphatase activity of the cells remained associated with the mureinoplasts when the outer layers of the cell wall were removed. Upon treatment of the mureinoplasts with lysozyme, some alkaline phosphatase was released into the medium and some remained with the protoplasts formed. Cells washed and suspended in 0.5 M NaCl were lysed by treatment with 2% toluene, and 95% of the alkaline phosphatase in the cells was released into the medium. Cells washed and suspended in complete salts solution (0.3 M NaCl, 0.05 M MgSO(4), and 0.01 M KCl) or 0.05 M MgSO(4) appeared intact after treatment with toluene but lost 50 and 10%, respectively, of their alkaline phosphatase. The results suggest that the presence of Mg(2+) in the cell wall is necessary to prevent disruption of the cells by toluene and may also be required to prevent the release of alkaline phosphatase by toluene when disruption of the cells by toluene does not take place. 相似文献
16.
Glutathione reductase was extracted from toluene-treated baker's yeast cells by a two-stage buffer autolysis method. The yeast cells were treated with toluene for 1 h at 40 degrees C. After removal of the toluene, the cells were then allowed to autolysis in buffer for 72 h at 4 degrees C. The cells were collected and resuspended in buffer. A second stage autolysis was carried out for another 96 h at 4 degrees C. The enzyme was purified to 786-fold from the second stage cell autolysate by using two steps of affinity chromatography with triazine dyes (Yellow H-E4G and Yellow H-E6G) coupled to Sepharose CL-4B. By using this simplified method, 1.44 mg (165 units/mg) of glutathione reductase was obtained from 65 g (wet weight) of yeast cells, equivalent to 80% enzyme recovery. 相似文献
17.
Michael Nüsse 《Cell proliferation》1982,15(5):529-543
Abstract. Ehrlich ascites tumour cells growing in vitro in suspension culture were separated according to volume by the technique of velocity sedimentation in a zonal rotor with a reorienting gradient. Using DNA distribution analysis the sedimentation pattern of the cells could be analysed in detail. With appropriate conditions it was possible to separate pure G1 cells. Samples could also be obtained which were enriched in S or G2 + M cells. The main limitation of the selection in this type of rotor was the reorientation of the gradient which caused disturbances during deceleration of the rotor. The synchronous growth of selected G1 cells has been studied in detail to investigate the reasons for the rather poor synchrony of these cells. The poor synchrony was found to be caused mainly by the small volume of the selected G1 cells compared with the normal volume of G1 cells in an asynchronous population. The synchronization of these cells could be essentially improved by a short treatment with excess thymidine causing a metabolic block at the G1/S border. The duration of this treatment could be minimized using DNA distribution analysis of growing cells after releasing of the block. The durations of the cell cycle phases in synchronized cells agreed with the values calculated in asynchronous cells by DNA distribution analysis and the BrdUrd-Hoechst 33258-technique. 相似文献
18.
19.
Kanter M 《Neurochemical research》2008,33(3):579-588
The aim of this study was designed to investigate the possible beneficial effects of Nigella sativa (NS) and derived thymoquinone (TQ) on neurodegeneration in hippocampus after chronic toluene exposure in rats. The rats were
randomly allotted into one of four experimental groups: A (control), B (toluene treated), C (toluene treated with NS) and
D (toluene treated with TQ); each group contain 10 animals. Toluene treatment was performed by inhalation of 3,000 ppm toluene,
in a 8 h/day and 6 day/week order for 12 weeks. Control group received 1 ml serum physiologic and the rats in NS and TQ treated
groups (C and D) were given NS (in a dose of 400 mg/kg body weight) and TQ (50 mg/kg body weight) once a day orally by using
intra gastric intubation for 12 weeks starting just after toluene exposure respectively. Tissue samples were obtained for
histopathological investigation. To date, no histopathological changes of neurodegeneration in hippocampus after chronic toluene
exposure in rats by NS and TQ treatment have been reported. In this study, chronic toluene exposure caused severe degenerative
changes, shrunken cytoplasma, slightly dilated cisternae of endoplasmic reticulum, markedly swollen mitochondria with degenerated
cristae and nuclear membrane breakdown with chromatin disorganization in neurons of the hippocampus. The distorted nerve cells
were mainly absent in the TQ and NS-treated rats. We conclude that TQ and especially NS therapy causes morphologic improvement
on neurodegeneration in hippocampus after chronic toluene exposure in rats. We believe that further preclinical research into
the utility of NS and TQ may indicate its usefulness as a potential treatment on neurodegeneration after chronic toluene exposure
in rats. 相似文献
20.
Jackson, Robert W. (University of California, San Diego, La Jolla), and J. A. DeMoss. Effects of toluene on Escherichia coli. J. Bacteriol. 90:1420-1425. 1965.-When toluene is added at appropriate levels to exponentially growing cultures of Escherichia coli, a time-dependent loss of turbidity is observed which is concurrent with a loss of material to the medium and with unmasking of beta-galactosidase. In addition, the galactoside permease system is totally destroyed. Electron micrographs confirm the indications that the cells are not being lysed by toluene, although the cytoplasm collapses to the interior of the cell. Included in the material lost from the cell after toluene treatment is 85% of the total ribonucleic acid (RNA), the principal source of which appears to be the ribosomes. The loss of RNA is temperature-dependent. Protein is also lost to the medium as a function of both temperature and available toluene. Up to 25% of the total protein is found in the medium, the precise amount depending on the level of toluene employed. Zone centrifugation studies of extracts from treated cells indicate that toluene elicits a rapid disaggregation of ribosomes that is terminated, at any stage, by disruption of the cells. The disaggregation is temperature-dependent and does not occur at 4 C. It appears to be distinct from the actual degradation of ribosomal RNA and is accompanied by an accumulation of small particles during the initial phases of treatment at 21 C. Toluene added to crude extracts of normal E. coli cells is unable to cause detectable ribosome destruction. 相似文献