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Pseudomonas putida and Pseudomonas fluorescens present as a coculture were studied for their abilities to degrade benzene, toluene, ethylbenzene, and xylenes (collectively known as BTEX) under various growth conditions. The coculture effectively degraded various concentrations of BTEX as sole carbon sources. However, all BTEX compounds showed substrate inhibition to the bacteria, in terms of specific growth, degradation rate, and cell net yield. Cell growth was completely inhibited at 500mgl–1 of benzene, 600mgl–1 of o-xylene, and 1000mgl–1 of toluene. Without aeration, aerobic biodegradation of BTEX required additional oxygen provided as hydrogen peroxide in the medium. Under hypoxic conditions, however, nitrate could be used as an alternative electron acceptor for BTEX biodegradation when oxygen was limited and denitrification took place in the culture. The carbon mass balance study confirmed that benzene and toluene were completely mineralized to CO2 and H2O without producing any identifiable intermediate metabolites. 相似文献
3.
The stability of Pseudomonas putida F1, a strain harbouring the genes responsible for toluene degradation in the chromosome was evaluated in a bioscrubber under
high toluene loadings and nitrogen limiting conditions at two dilution rates (0.11 and 0.27 h−1). Each experiment was run for 30 days, period long enough for microbial instability to occur considering previously reported
studies carried out with bacterial strains encoding the catabolic genes in the TOL plasmid. At all tested conditions, P. putida F1 exhibited stable performance as shown by the constant values of the specific toluene degradation yield, CO2 produced versus toluene degraded yield, and biomass concentration within each steady state. Benzyl alcohol, a curing agent
causing TOL plasmid deletion in Pseudomonas strains, was present in the cultivation medium as a result of the monooxygenation of toluene by the diooxygenase system of
P. putida F1. However, no mutant population growing at the expense of the extracellular excreted carbon or lysis products was established
in the chemostat as confirmed by the constant dissolved total organic carbon (TOC) concentration and fraction of toluene degrading
cells (approx. 100%). In addition, batch experiments conducted with both lysis substrate and toluene simultaneously confirmed
that P. putida F1 preferentially consumed toluene rather than extracellular excreted carbon. 相似文献
4.
Biotransformation of isoeugenol to vanillin by <Emphasis Type="Italic">Pseudomonas putida</Emphasis> IE27 cells 总被引:1,自引:0,他引:1
The ability to produce vanillin and/or vanillic acid from isoeugenol was screened using resting cells of various bacteria.
The vanillin- and/or vanillic-acid-producing activities were observed in strains belonging to the genera Achromobacter, Aeromonas, Agrobacerium, Alcaligenes, Arthrobacter, Bacillus, Micrococcus, Pseudomonas, Rhodobacter, and Rhodococcus. Strain IE27, a soil isolate showing the highest vanillin-producing activity, was identified as Pseudomonas putida. We optimized the culture and reaction conditions for vanillin production from isoeugenol using P. putida IE27 cells. The vanillin-producing activity was induced by adding isoeugenol to the culture medium but not vanillin or eugenol.
Under the optimized reaction conditions, P. putida IE27 cells produced 16.1 g/l vanillin from 150 mM isoeugenol, with a molar conversion yield of 71% at 20 °C after a 24-h
incubation in the presence of 10% (v/v) dimethyl sulfoxide. 相似文献
5.
The fadBA operon in the fatty acid β-oxidation pathway of P. putida KCTC1639 was blocked to induce a metabolic flux of the intermediates to the biosynthesis of medium chain-length PHA (mcl-PHA).
Succinate at 150 mg l−1 stimulated cell growth and also the biosynthesis of medium chain-length-polyhydroxyalkanoate. pH-stat fed-batch cultivation
of the fadA knockout mutant P. putida KCTC1639 was carried out for 60 h, in which mcl-PHA reached 8 g l−1 with a cell dry weight of 10.3 g l−1. 相似文献
6.
The response of Pseudomonas putida F1 to process fluctuations and operational failures during toluene biodegradation was evaluated in a chemostat suspended
growth bioreactor. The ability of P. putida F1 to rapidly increase its specific toluene degradation capacity resulted in no significant variation in process removal
efficiency when toluene load was increased from 188 to 341 g m−3 h−1. Likewise, bacterial activity rapidly reached steady state performance (in less than 1.5 h after the restoration of steady
state operational conditions) following an 8 h process shutdown, or after episodes of toluene or mineral nutrients deprivation.
Process performance was however highly sensitive to pH, as pH levels below 4.5 dramatically inhibited bacterial activity,
decreasing severely process robustness and inducing a cycle of periodic process collapses and recoveries. This pH mediated
deterioration of bacterial activity was confirmed by further respirometric tests, which revealed a 50–60% reduction in the
O2 consumption rate during the degradation of both toluene and 3-methyl catechol when pH decreased from 5.05 to 4.55. Finally,
process robustness was quantified according to methods previously described in literature. 相似文献
7.
Pseudomonas putida KT2440 grew on glucose at a specific rate of 0.48 h−1 but accumulated almost no poly-3-hydroxyalkanoates (PHA). Subsequent nitrogen limitation on nonanoic acid resulted in the
accumulation of only 27% medium-chain-length PHA (MCL-PHA). In contrast, exponential nonanoic acid-limited growth (μ = 0.15 h−1) produced 70 g l−1 biomass containing 75% PHA. At a higher exponential feed rate (μ = 0.25 h−1), the overall productivity was increased but less biomass (56 g l−1) was produced due to higher oxygen demand, and the biomass contained less PHA (67%). It was concluded that carbon-limited
exponential feeding of nonanoic acid or related substrates to cultures of P. putida KT2440 is a simple and highly effective method of producing MCL-PHA. Nitrogen limitation is unnecessary. 相似文献
8.
Pseudomonas putida CP1 formed clumps of cells when grown on mono-chlorophenols but not on phenol or glucose. An increase in cell numbers for
the organism grown on mono-chlorophenols was accompanied by a decrease in the dry weight. The change in shape of the bacterium
from rod shape to coccus shape coupled with a reduction in cell size when the organism was grown under nutritional stress
was found. This result together with cell aggregation affected the measurement of growth parameters in the system by conventional
methods (optical density measurements, dry weight measurements and the plate count technique). Monitoring growth of Pseudomonas putida CP1 by a direct microscopic count technique was found to be more representative than conventional methods including optical
density measurements, dry weight measurements and the plate count technique when grown on phenolics. 相似文献
9.
Zhiyong Sun Juliana A. Ramsay Martin Guay Bruce A. Ramsay 《Applied microbiology and biotechnology》2009,82(4):657-662
Unsaturated medium-chain-length polyhydroxyalkanoates (MCL-PHA) were produced at a productivity of 0.63–1.09 g PHA l−1 h−1 with final PHA content ranging from 42.6 to 55.8% in single-stage, carbon-limited, fed-batch fermentations of Pseudomonas putida KT2440. A mixture of nonanoic acid (NA) and 10-undecenoic acid (UDA=) was fed exponentially to control growth rate. Varying the specific growth rate (0.14 h−1 vs. 0.23 h−1) at similar substrate feed ratios (NA:UDA= = 5:1) had little effect on the final PHA content and relative composition. However, decreasing the NA:UDA= ratio decreased the final amount of PHA produced from 56% with NA:UDA= = 5.07:1 to only 42% at NA:UDA= = 2.47:1. The molar fraction of all 3-hydroxyalkanoate monomers in the PHA product was relatively constant throughout each
fermentation, indicating that the final product was homogeneous rather than a mixture of different copolymers. A linear relationship
between unsaturation of the PHA produced and unsaturation of the carbon feed was found, which demonstrates the feasibility
of producing unsaturated MCL-PHAs with controlled polymeric composition in a fed-batch process. 相似文献
10.
Arene cis-diols are interesting chemicals because of their chiral structures and great potentials in industrial synthesis of useful
chiral chemical products. Pseudomonas putida KT2442 was genetically modified to transform benzoic acid (benzoate) to 1,2-dihydroxy-cyclohexa-3,5-diene-1-carboxylic acid
(DHCDC) or named benzoate cis-diol. BenD gene encoding cis-diol dehydrogenase was deleted to generate a mutant named P. putida KTSY01. Genes benABC encoding benzoate dioxygenase were cloned into plasmid pSYM01 and overexpressed in P. putida KTSY01. The recombinant bacteria P. putida KTSY01 (pSYM01) showed strong ability to transform benzoate to DHCDC. DHCDC of 2.3 g/L was obtained with a yield of 73% after
24 h of cultivation in shake flasks incubated under optimized growth conditions. Transformation of benzoate carried out in
a 6-L fermentor using a benzoate fed-batch process produced over 17 g/L DHCDC after 48 h of fermentation. The average DHCDC
production rate was 0.356 g L−1 h−1. DHCDC purified from the fermentation broth showed a purity of more than 95%, and its chemical structure was confirmed by
nuclear magnetic resonance. 相似文献
11.
Singh AK Chaudhary P Macwan AS Diwedi UN Kumar A 《Applied microbiology and biotechnology》2007,76(4):895-901
The chlorinated insecticide γ-hexachlorocyclohexane (γ-HCH) is sequentially metabolized by the products of linA, linB, linC, linD, linE, and linF genes to β-ketoadipate, which is subsequently mineralized. Two or more copies of these genes are present in the bacterium
Pseudomonas aeruginosa ITRC-5 that was isolated earlier by selective enrichment on technical-HCH. At least one copy of linA, linB, linC, linD, and possibly linE is lost from ITRC-5 upon its growth on γ-HCH. All the lin genes, however, are lost when the bacterium was grown in Luria–Bertani (LB) medium. The loss of lin genes is accompanied with the loss/rearrangement of insertion sequence IS6100 genes. Concomitant to the loss of lin genes, the degradation of HCH-isomers by “γ-HCH grown cells” is slower, when compared with “technical-HCH grown cells”, and
is completely lost by “LB-grown cells”. The selective loss of lin genes during different growth conditions has not been reported before and is expected to help in understanding the dynamism
of degradative genes. 相似文献
12.
The gene encoding a Baeyer-Villiger monooxygenase and identified in Pseudomonas putida KT2440 was cloned and functionally expressed in Escherichia coli. The highest yield of soluble protein could be achieved by co-expression of molecular chaperones. In order to determine the
substrate specificity, biocatalyses were performed using crude cell extract, growing and resting cells. Examination of aromatic,
cyclic and aliphatic ketones revealed a high specificity towards short-chain aliphatic ketones. Interestingly, some open-chain
ketones were converted to the alkylacetates, while for others formation of the ester products with oxygen on the other side
of the keto group could also be detected yielding the corresponding methyl or ethyl esters. 相似文献
13.
Purification,characterization and gene cloning of isoeugenol-degrading enzyme from <Emphasis Type="Italic">Pseudomonas putida</Emphasis> IE27 总被引:1,自引:0,他引:1
An isoeugenol-degrading enzyme was purified to homogeneity from Pseudomonas putida IE27, an isoeugenol-assimilating bacterium. The purified enzyme was a 55 kDa monomer and catalyzed the initial step of isoeugenol
degradation, the oxidative cleavage of the side chain double-bond of isoeugenol, to form vanillin. Another reaction product
of isoeugenol degradation besides vanillin was identified to be acetaldehyde. The values of Km and k
cat for isoeugenol were 175 μM and 5.18 s–1, respectively. The purified enzyme catalyzed the incorporation of an oxygen atom from either molecular oxygen or water into
vanillin, suggesting that the isoeugenol-degrading enzyme is a kind of monooxygenase. The gene encoding the isoeugenol-degrading
enzyme and its flanking regions were isolated from P. putida IE27. The amino acid sequence of the enzyme was similar to those of lignostilbene-α,β-dioxygenases, carotenoid monooxygenases
and 9-cis-epoxycarotenoid dioxygenases. 相似文献
14.
Pseudomonas putida CSV86 utilizes naphthalene (Nap), salicylate (Sal), benzyl alcohol (Balc), and methylnaphthalene (MN) preferentially over
glucose. Methylnaphthalene is metabolized by ring-hydroxylation as well as side-chain hydroxylation pathway. Although the
degradation property was found to be stable, the frequency of obtaining Nap−Sal−MN−Balc− phenotype increased to 11% in the presence of curing agents. This property was transferred by conjugation to Stenotrophomonas maltophilia CSV89 with a frequency of 7 × 10−8 per donor cells. Transconjugants were Nap+Sal+MN+Balc+ and metabolized MN by ring- as well as side-chain hydroxylation pathway. Transconjugants also showed the preferential utilization
of aromatic compounds over glucose indicating transfer of the preferential degradation property. The transferred properties
were lost completely when transconjugants were grown on glucose or 2YT. Attempts to detect and isolate plasmid DNA from CSV86
and transconjugants were unsuccessful. Transfer of degradation genes and its subsequent loss from the transconjugants was
confirmed by PCR using primers specific for 1,2-dihydroxynaphthalene dioxygenase and catechol 2,3-dioxygenase (C23O) as well
as by DNA–DNA hybridizations using total DNA as template and C23O PCR fragment as a probe. These results indicate the involvement
of a probable conjugative element in the: (i) metabolism of aromatic compounds, (ii) ring- and side-chain hydroxylation pathways
for MN, and (iii) preferential utilization of aromatics over glucose. 相似文献
15.
Toluene dioxygenase (TDO) catalyzes asymmetric cis-dihydroxylation of aromatic compounds. To achieve high efficient biotransformation of benzene to benzene cis-diols, Pseudomonas putida KT2442, Pseudomonas stutzeri 1317, and Aeromonas hydrophila 4AK4 were used as hosts to express TDO gene tod. Plasmid pSPM01, a derivative of broad-host plasmid pBBR1MCS-2 harboring tod from plasmid pKST11, was constructed and introduced into the above three strains. Their abilities to catalyze the biotransformation
of benzene to benzene cis-diols, namely, cis-3,5-cyclohexadien-1,2-diols abbreviated as DHCD, were examined. In shake-flask cultivation under optimized culture media
and growth condition, benzene cis-diols production by recombinant P. putida KT2442 (pSPM01), P. stutzeri 1317 (pSPM01), and A. hydrophila 4AK4 (pSPM01) were 2.68, 2.13, and 1.17 g/l, respectively. In comparison, Escherichia coli JM109 (pSPM01) and E. coli JM109 (pKST11) produced 0.45 and 0.53 g/l of DHCD, respectively. When biotransformation was run in a 6-l fermenter, DHCD
production in P. putida KT2442 (pSPM01) was approximately 60 g/l; this is the highest DHCD production yield reported so far. 相似文献
16.
Chen X Shi J Chen Y Xu X Chen L Wang H Hu T 《Applied microbiology and biotechnology》2007,74(4):881-889
Previously performed studies have shown that Pseudomonas putida CZ1 biomass can bind an appreciable amount of Cu(II) and Zn(II) ions from aqueous solutions. The mechanisms of Cu- and Zn-binding
by P. putida CZ1 were ascertained by chemical modifications of the biomass followed by Fourier transform infrared and X-ray absorption
spectroscopic analyses of the living or nonliving cells. A dramatic decrease in Cu(II)- and Zn(II)-binding resulted after
acidic methanol esterification of the nonliving cells, indicating that carboxyl functional groups play an important role in
the binding of metal to the biomaterial. X-ray absorption spectroscopy was used to determine the speciation of Cu ions bound
by living and nonliving cells, as well as to elucidate which functional groups were involved in binding of the Cu ions. The
X-ray absorption near-edge structure spectra analysis showed that the majority of the Cu was bound in both samples as Cu(II).
The fitting results of Cu K-edge extended X-ray absorption fine structure spectra showed that N/O ligands dominated in living
and nonliving cells. Therefore, by combining different techniques, our results indicate that carboxyl functional groups are
the major ligands responsible for the metal binding in P. putida CZ1. 相似文献
17.
Banerjee A Dubey S Kaul P Barse B Piotrowski M Banerjee UC 《Molecular biotechnology》2009,41(1):35-41
Nitrilases have attracted tremendous attention for the preparation of optically pure carboxylic acids. This article aims to
address the production and utilization of a highly enantioselective nitrilase from Pseudomonas putida MTCC 5110 for the hydrolysis of racemic mandelonitrile to (R)-mandelic acid. The nitrilase gene from P. putida was cloned in pET 21b(+) and over-expressed as histidine-tagged protein in Escherichia coli. The histidine-tagged enzyme was purified from crude cell extracts of IPTG-induced cells of E. coli BL21 (DE3). Inducer replacement studies led to the identification of lactose as a suitable and cheap alternative to the costly
IPTG. Effects of medium components, various physico-chemical, and process parameters (pH, temperature, aeration, and agitation)
for the production of nitrilase by engineered E. coli were optimized and scaled up to a laboratory scale bioreactor (6.6 l). Finally, the recombinant E. coli whole-cells were utilized for the production of (R)-(−)-mandelic acid. 相似文献
18.
A new isolate of <Emphasis Type="Italic">Pseudomonas stutzeri</Emphasis>that degrades 2-chloronitrobenzene 总被引:3,自引:0,他引:3
A strain of Pseudomonas stutzeri ZWLR2-1 was isolated from soil contaminated with chloronitrobenzenes and identified by 16S rDNA sequencing. This bacterium released chloride and nitrite into the medium when grown on 0.5mm 2-chloronitrobenzene. PCR amplification and DNA sequencing revealed a DNA fragment encoding a polypeptide homologous to the -subunit of ring-hydroxylating dioxygenasesRevisions requested 01 December 2004; Revisions received 9 December 2004 相似文献
19.
The maT clade of transposons is a group of transposable elements intermediate in sequence and predicted protein structure to mariner and Tc transposons, with a distribution thus far limited to a few invertebrate species. We present evidence, based on searches of publicly available databases, that the nematode Caenorhabditis briggsae has several maT-like transposons, which we have designated as CbmaT elements, dispersed throughout its genome. We also describe two additional transposon sequences that probably share their evolutionary history with the CbmaT transposons. One resembles a fold back variant of a CbmaT element, with long (380-bp) inverted terminal repeats (ITRs) that show a high degree (71%) of identity to CbmaT1. The other, which shares only the 26-bp ITR sequences with one of the CbmaT variants, is present in eight nearly identical copies, but does not have a transposase gene and may therefore be cross mobilised by a CbmaT transposase. Using PCR-based mobility assays, we show that CbmaT1 transposons are capable of excising from the C. briggsae genome. CbmaT1 excised approximately 500 times less frequently than Tcb1 in the reference strain AF16, but both CbmaT1 and Tcb1 excised at extremely high frequencies in the HK105 strain. The HK105 strain also exhibited a high frequency of spontaneous induction of unc-22 mutants, suggesting that it may be a mutator strain of C. briggsae. 相似文献
20.
Fusarium solani degraded methyl tert-butyl ether (MTBE) and other oxygenated compounds from gasoline including tert-butyl alcohol (TBA). The maximum degradation rate of MTBE was 16 mg protein h and 46 mg/g protein h for TBA. The culture
transformed 77% of the total carbon to 14CO2. The estimated yield for MTBE was 0.18 g dry wt/g MTBE. 相似文献