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1.
Forty-six Rhizobium isolates from legume root and stem nodules were examined for their phosphate-solubilizing ability on Pikovskaya’s agar medium.
Rhizobium isolates from root nodules of Cassia absus, Vigna trilobata and three strains from Sesbania sesban showed zone of tricalcium phosphate (TCP) solubilization. The isolate from C. absus showed maximum solubilization (620 μg/ml) after 12 d of incubation, while the Rhizobium sp. strain 26 (from S. sesban) showed the least amount (150 μg/ml) of phosphate solubilization. Among the carbon sources tested for their ability to solubilize
TCP, maximum solubilization (620 μg/ml) was observed in glucose by Rhizobium isolate from C. absus. Phosphate solubilization increased with increase in glucose concentration steeply up to 2% and slowly
above this concentration in four isolates. Among the nitrogen sources tested, maximum solubilization (620 μg/ml) was observed
in ammonium sulphate by Rhizobium isolate from C. absus. 相似文献
2.
Chanchal Kumar Jitendra Wagh G. Archana G. Naresh Kumar 《World journal of microbiology & biotechnology》2016,32(12):194
Enterobacter asburiae PSI3 solubilizes mineral phosphates in the presence of glucose by the secretion of gluconic acid generated by the action of a periplasmic pyrroloquinoline quinone dependent glucose dehydrogenase. In order to achieve mineral phosphate solubilization phenotype in the presence of sucrose, plasmids pCNK4 and pCNK5 containing genes encoding the invertase enzyme of Zymomonas mobilis (invB) and of Saccharomyces cerevisiae (suc2) under constitutive promoters were constructed with malE signal sequence (in case of invB alone as the suc2 is secreted natively). When introduced into E. asburiae PSI3, E. a. (pCNK4) and E. a. (pCNK5) transformants secreted 21.65 ± 0.94 and 22 ± 1.3 mM gluconic acid, respectively, in the presence of 75 mM sucrose and they also solubilized 180 ± 4.3 and 438 ± 7.3 µM P from the rock phosphate. In the presence of a mixture of 50 mM sucrose and 25 mM glucose, E. a. (pCNK5) secreted 34 ± 2.3 mM gluconic acid and released 479 ± 8.1 µM P. Moreover, in the presence of a mixture of eight sugars (10 mM each) in the medium, E. a. (pCNK5) released 414 ± 5.3 µM P in the buffered medium. Thus, this study demonstrates incorporation of periplasmic invertase imparted P solubilization ability to E. asburiae PSI3 in the presence of sucrose and mixture of sugars. 相似文献
3.
The nifA gene is an important regulatory gene and its product, NifA protein, regulates the expression of many nif genes involved in the nitrogen fixation process. We introduced multiple copies of the constitutively expressed Sinorhizobium meliloti (Sm) or Enterobacter cloacae (Ec) nifA gene into both the nifA mutant strain SmY and the wild-type strain Sm1021. Root nodules produced by SmY containing a constitutively expressed Sm nifA gene were capable of fixing nitrogen, while nodules produced by SmY containing the Ec nifA gene remained unable to fix nitrogen, as is the case for SmY itself. However, transfer of an additional Sm nifA gene into Sm1021 improved the nitrogen-fixing efficiency of root nodules to a greater extent than that observed upon transfer
of the Ec nifA gene into Sm1021. Comparative analysis of amino acid sequences between Sm NifA and Ec NifA showed that the N-terminal domain was the least similar, but this domain is indispensable for complementation of the
Fix- phenotype of SmY by Sm NifA. We conclude that more than one domain is involved in determining functional differences between Sm NifA and Ec NifA. 相似文献
4.
Mahesh S. Yandigeri Arvind K. Yadav R. Srinivasan Sudhanshu Kashyap Sunil Pabbi 《Microbiology》2011,80(4):558-565
Two diazotrophic cyanobacteria, Westiellopsis prolifica and Anabaena variabilis were evaluated for elucidating the possible mechanism of mineral phosphate solubilization. Phosphate starved cyanobacteria
evaluated for the presence of organic acids, extracellular compounds or enzymes that might have been produced and promoted
the mineral phosphate solubilization with Mussorie Rock Phosphate and Tricalcium Phosphate as substrates. Both the cultures
did not reveal production of organic acids throughout the incubation period when checked for decrease in media pH of the media
and thin layer chromatography. Thin layer chromatography of culture filtrates showed the presence of hydrocarbon like compound.
Further analysis of the culture filtrates with gas liquid chromatography, a single peak near to the retention time of 7.6
was observed in all extracts of culture filtrates irrespective of phosphate source. UV-visible spectra of culture filtrates
revealed the absorption maxima of 276 nm. Gas chromatographic-mass spectrometric analysis of culture filtrates showed most
intense peak in the electron impact (EI) ionization was at m/z 149 and molecular ion peaks at m/z 207 and 167, inferring the presence of phthalic acid. Among the mechanisms in mineral phosphate solubilization, it was evident
that these cyanobacteria used phthalic acid as possible mode of P solubilization. 相似文献
5.
Livingstone JR Yoshida I Tarui Y Hirooka K Yamamoto Y Tsutui N Hirasawa E 《Journal of plant research》2002,115(5):393-400
NAD-dependent aminoaldehyde dehydrogenase (AMADH, EC 1.2.1.-) from Avena shoots was purified by DEAE Sephacel, hydroxyapatite, 5′-AMP Sepharose 4B, Mono Q, and TSK-GEL column chromatographies to
homogeneity by the criterion of native PAGE. SDS–PAGE yielded a single band at a molecular mass of 55 kDa. IEF studies showed
a band with a pI value of 5.3. In contrast to AMADHs from other species, the TSK-GEL chromatography showed that Avena AMADH exists as a monomer in the native state. The purified enzyme catalyzed the oxidations of 3-aminopropionaldehyde (APAL),
4-aminobutyraldehyde (ABAL) N-(3-aminopropyl)-4-aminobutyraldehyde (APBAL), and 4-guanidinobutyraldehyde (GBAL), but not of betaine aldehyde or indoleacetaldehyde.
The K
m values for APAL, ABAL, and GBAL were 1.5×10–6, 2.2×10–6, and 1.3×10–5 M, respectively. Although N-terminal amino acid sequence of Avena AMADH could not be determined due to a modification of the amino residue, the sequence of the fragment of AMADH cleaved by
V8 protease showed greater similarity to the barley BADH than to the pea AMADH.
Electronic Publication 相似文献
6.
A FAD-dependent glucose dehydrogenase (FADGDH) mutant with narrow substrate specificity was constructed by site-directed mutagenesis.
Several characteristics of FADGDH, such as high catalytic activity and high electron transfer ability, make this enzyme suitable
for application to glucose sensors. However, for further applications, improvement of the broad substrate specificity is needed.
In this paper, we mutated two residues, Asn475 and Ala472, which are located near the putative active site of the catalytic
subunit of FADGDH and have been predicted from the alignment with the active site of glucose oxidase. Of the 38 mutants constructed,
Ala472Phe and Asn475Asp were purified and their activities were analyzed. Both mutants showed a higher specificity toward
glucose compared to the wild type enzyme. 相似文献
7.
Enterobacter aerogenes NBRC12010 was able to ferment glycerol to ethanol and hydrogen gas. Fermentation of glycerol ceased in the stationary phase
of growth, and it was activated by electrochemical reactions using thionine as an electron transfer mediator from bacterial
cells to an electrode. Using resting cells of E. aerogenes NBRC12010 in only citrate buffer solution, the cells did not consume glycerol at all, but they could metabolize glucose.
These results suggest that the regulation of glycerol metabolism occurred at enzymatic steps before glycolysis. In E. aerogenes NBRC12010, glycerol was metabolized via glycerol dehydrogenase (GDH) and then dehydroxyacetone kinase. The GDH-catalyzed
reaction mainly depended on the ratio of NAD+/NADH. At a NAD+/NADH ratio of nearly 1 or less, it was substantially suppressed and glycerol metabolism stopped. When the ratio was higher
than 1, GDH was activated and glycerol was metabolized. Thus, the reaction of glycerol metabolism depended on the balance
of cellular NAD+/NADH. Exogenous NADH was oxidized to NAD+ by electrochemical reactions with thionine. We proposed the activation mechanism of glycerol metabolism under electrochemical
conditions. 相似文献
8.
Ma C Gao C Qiu J Hao J Liu W Wang A Zhang Y Wang M Xu P 《Applied microbiology and biotechnology》2007,77(1):91-98
Pseudomonas stutzeri SDM was newly isolated from soil, and two stereospecific NAD-independent lactate dehydrogenase (iLDH) activities were detected
in membrane of the cells cultured in a medium containing dl-lactate as the sole carbon source. Neither enzyme activities was constitutive, but both of them might be induced by either
enantiomer of lactate. P. stutzeri SDM preferred to utilize lactate to growth, when both l-lactate and glucose were available, and the consumption of glucose was observed only after lactate had been exhausted. The
Michaelis–Menten constant for l-lactate was higher than that for d-lactate. The l-iLDH activity was more stable at 55°C, while the d-iLDH activity was lost. Both enzymes exhibited different solubilization with different detergents and different oxidation
rates with different electron acceptors. Combining activity staining and previous proteomic analysis, the results suggest
that there are two separate enzymes in P. stutzeri SDM, which play an important role in converting lactate to pyruvate.
Ma and Gao contributed equally to this work. 相似文献
9.
Skory CD 《Journal of industrial microbiology & biotechnology》2003,30(1):22-27
This work demonstrates the first example of a fungal lactate dehydrogenase (LDH) expressed in yeast. A L(+)-LDH gene, ldhA, from the filamentous fungus Rhizopus oryzae was modified to be expressed under control of the Saccharomyces cerevisiae adh1 promoter and terminator and then placed in a 2μ-containing yeast-replicating plasmid. The resulting construct, pLdhA68X,
was transformed and tested by fermentation analyses in haploid and diploid yeast containing similar genetic backgrounds. Both
recombinant strains utilized 92 g glucose/l in approximately 30 h. The diploid isolate accumulated approximately 40% more
lactic acid with a final concentration of 38 g lactic acid/l and a yield of 0.44 g lactic acid/g glucose. The optimal pH for
lactic acid production by the diploid strain was pH 5. LDH activity in this strain remained relatively constant at 1.5 units/mg
protein throughout the fermentation. The majority of carbon was still diverted to the ethanol fermentation pathway, as indicated
by ethanol yields between 0.25–0.33 g/g glucose. S. cerevisiae mutants impaired in ethanol production were transformed with pLdhA68X in an attempt to increase the lactic acid yield by
minimizing the conversion of pyruvate to ethanol. Mutants with diminished pyruvate decarboxylase activity and mutants with
disrupted alcohol dehydrogenase activity did result in transformants with diminished ethanol production. However, the efficiency
of lactic acid production also decreased.
Electronic Publication 相似文献
10.
Vanessa Sayuri Sato Renato F. Galdiano Júnior Gisele Regina Rodrigues Eliana G. M. Lemos João Martins Pizauro Junior 《Journal of microbiology (Seoul, Korea)》2016,54(2):106-113
Expression of acid ectophosphatase by Enterobacter asburiae, isolated from Cattleya walkeriana (Orchidaceae) roots and identified by the 16S rRNA gene sequencing analysis, was strictly regulated by phosphorus ions, with its optimal activity being observed at an inorganic phosphate concentration of 7 mM. At the optimum pH 3.5, intact cells released p-nitrophenol at a rate of 350.76 ± 13.53 nmol of p-nitrophenolate (pNP)/min/108 cells. The membrane-bound enzyme was obtained by centrifugation at 100,000 × g for 1 h at 4°C. p-Nitrophenylphosphate (pNPP) hydrolysis by the enzyme follows “Michaelis-Menten” kinetics with V = 61.2 U/mg and K0.5 = 60 μM, while ATP hydrolysis showed V = 19.7 U/mg, K0.5 = 110 μM, and nH = 1.6 and pyrophosphate hydrolysis showed V = 29.7 U/mg, K0.5 = 84 μM, and nH = 2.3. Arsenate and phosphate were competitive inhibitors with Ki = 0.6 mM and Ki = 1.8 mM, respectively. p-Nitrophenyl phosphatase (pNPPase) activity was inhibited by vanadate, while p-hydroxymercuribenzoate, EDTA, calcium, copper, and cobalt had no inhibitory effects. Magnesium ions were stimulatory (K0.5 = 2.2 mM and nH = 0.5). Production of an acid ectophosphatase can be a mechanism for the solubilization of mineral phosphates by microorganisms such as Enterobacter asburiae that are versatile in the solubilization of insoluble minerals, which, in turn, increases the availability of nutrients for plants, particularly in soils that are poor in phosphorus. 相似文献
11.
The gene encoding malate dehydrogenase (MDH) was overexpressed in a pflB ldhA double mutant of Escherichia coli, NZN111, for succinic acid production. With MDH overexpression, NZN111/pTrc99A-mdh restored the ability to metabolize glucose anaerobically and 0.55 g/L of succinic acid was produced from 3 g/L of glucose
in shake flask culture. When supplied with 10 g/L of sodium bicarbonate (NaHCO3), the succinic acid yield of NZN111/pTrc99A-mdh reached 1.14 mol/mol glucose. Supply of NaHCO3 also improved succinic acid production by the control strain, NZN111/pTrc99A. Measurement of key enzymes activities revealed
that phosphoenolpyruvate (PEP) carboxykinase and PEP carboxylase in addition to MDH played important roles. Two-stage culture
of NZN111/pTrc99A-mdh was carried out in a 5-L bioreactor and 12.2 g/L of succinic acid were produced from 15.6 g/L of glucose. Fed-batch culture
was also performed, and the succinic acid concentration reached 31.9 g/L with a yield of 1.19 mol/mol glucose. 相似文献
12.
<Emphasis Type="Italic">Thermotoga maritima</Emphasis> TM0298 is a highly thermostable mannitol dehydrogenase 总被引:1,自引:1,他引:0
Song SH Ahluwalia N Leduc Y Delbaere LT Vieille C 《Applied microbiology and biotechnology》2008,81(3):485-495
Thermotoga maritima TM0298 is annotated as an alcohol dehydrogenase, yet it shows high identity and similarity to mesophilic mannitol dehydrogenases.
To investigate this enzyme further, its gene was cloned and expressed in Escherichia coli. The purified recombinant enzyme was most active on fructose and mannitol, making it the first known hyperthermophilic mannitol
dehydrogenase. T. maritima mannitol dehydrogenase (TmMtDH) is optimally active between 90 and 100 °C and retains 63% of its activity at 120 °C but shows
no detectable activity at room temperature. Its kinetic inactivation follows a first-order mechanism, with half-lives of 57 min
at 80 °C and 6 min at 95 °C. Although TmMtDH has a higher V
max with NADPH than with NADH, its catalytic efficiency is 2.2 times higher with NADH than with NADPH and 33 times higher with
NAD+ than with NADP+. This cofactor specificity can be explained by the high density of negatively charged residues (Glu193, Asp195, and Glu196)
downstream of the NAD(P) interaction site, the glycine motif. We demonstrate that TmMtDH contains a single catalytic zinc
per subunit. Finally, we provide the first proof of concept that mannitol can be produced directly from glucose in a two-step
enzymatic process, using a Thermotoga neapolitana xylose isomerase mutant and TmMtDH at 60 °C. 相似文献
13.
14.
Kotrbova-Kozak A Kotrba P Inui M Sajdok J Yukawa H 《Applied microbiology and biotechnology》2007,76(6):1347-1356
15.
Appanna Vikram Ajjanna R. Alagawadi P. U. Krishnaraj K. S. Mahesh Kumar 《World journal of microbiology & biotechnology》2007,23(9):1333-1337
It has been previously shown that certain gram-negative bacteria do not have the ability to solubilize insoluble phosphates
due to the lack of pyrroloquinoline-quinone synthesis genes (pqq). PQQ is required as cofactor for the assembly of the glucose dehydrogenase (GDH) holoenzyme, which acts in the oxidation
of glucose to gluconic acid. In this context the transconjugation and expression of pqq genes in Azospirillum sp. was studied using the construct pMCG 898. pMCG 898 containing pqq gene/s was mobilized into an Azospirillum strain negative to mineral phosphate solubilization by biparental mating. The presence of the construct was also confirmed
by minipreps of the transconjugants. The transconjugants were able to solubilize dicalcium phosphate while the wild type was
not able to do so. The nitrogen-fixing ability of the transconjugants was also examined and they retained the ability to fix
nitrogen. Further detailed studies are required to confirm the utility of such strains in releasing inorganic P from fixed
phosphates in soil. 相似文献
16.
Samantha?Caixeta?de?Oliveira Gilberto?de?Oliveira Mendes Ubiana?Cássia?da?Silva Ivo?Ribeiro?da?Silva José?Ivo?Ribeiro?Júnior Maurício?Dutra?Costa
Microbial solubilization of rock phosphate (RP) is mainly achieved by the production of organic acids and medium acidification through H+ release. During RP solubilization, mineral nutrient availability is likely to be critical for determining how much carbon is channeled either for metabolite synthesis or for microbial growth, influencing organic acid release by microorganisms. Thus, the objective of this work was to study the relationships between the concentration of mineral nutrients in the growth medium and the efficiency of RP solubilization by Aspergillus niger FS1. For this, the fungus was grown in Czapek medium containing 0, 1, 2, 10, 50, and 100 % of its original concentration of mineral nutrients. Decreasing mineral availability in the growth medium led to decreases in fungal biomass and solubilized P, and increases in titratable acidity and solubilization efficiency as expressed by mg solubilized P per g fungal biomass (YP/B), indicating a shift in fungal metabolism from biomass production to organic acid release. The transfer of pre-grown biomass to media with or without added minerals confirmed that lower mineral availability increases YP/B and led to the solubilization of 76 % of P present in Patos RP. These observations open new perspectives on improving RP solubilization systems by manipulating mineral nutrient availability in the medium, with consequent gains in cost reduction. 相似文献
17.
A psychrotolerant phosphate solubilizing fungus has been isolated from the rock soil of a cold desert site in Indian Himalaya.
The fungus grows from 4 to 35°C (optimum 21°C), and from 2 to 13.5 pH (optimum 9) under laboratory conditions. Based on phenotypic
characters and 26S rDNA analysis, the fungus is identified as Paecilomyces hepiali. In quantitative estimation that was carried out at 9, 14, and 21°C, the fungus solubilized maximum phosphate at 14°C. In
view of the slow growth and persistence of the desired activity at low temperature, the estimation was carried out for a longer
period, i.e., up to 6 weeks. The suboptimal conditions for growth and biomass production were found to be optimal for phosphate
solubilization by the fungus. At 14 and 9°C, the solubilization touched its maximum on day 42. Decline in pH was found to
be significantly correlated with the phosphate solubilization at all the temperatures, under consideration. The acid phosphatase
activity was found to be more prominent than alkaline phosphatase in culture filtrate. High performance thin layer chromatography
(HPTLC) analysis showed production of six organic acids, gluconic and α-keto glutaric acid being in maximum amount in the
culture filtrate. The study has ecological significance in view of the nutrient cycling under low temperature environment,
prevalent in Himalayan region. 相似文献
18.
Ling Zhao Yongming Bao Jingyun Wang Boshi Liu Lijia An 《World journal of microbiology & biotechnology》2009,25(1):57-64
A mutant designated as UV-3 was obtained from wild-type Enterobacter aerogenes 10293 through u.v. radiation. The activities of α-acetolactate decarboxylase (Ald), lactate dehydrogenase (Ldh) and diacetyl
reductase (Dr) in UV-3 were strongly attenuated, with the lowest activities at pH 7.0–7.5, and temperature between 36 and
39°C. Compared to the wild-type, the yield of diacetyl by UV-3 was increased 18.7-fold, up to 1.05 ± 0.01 g l−1. Acetoin and ethanol productions were decreased by 48.4 and 71.4%, respectively, but acetate yield was increased by 34.6%.
Optimum medium for diacetyl production by UV-3 contained 10% glucose, 0.5% peptone, 0.5% yeast extract powder, 0.01% (NH4)2SO4, 0.1% citric acid, 0.2% MnSO4 and 0.2% MgSO4, and this was determined by one-factor-at-a-time approach. Data from the five level central composite designs demonstrated
that initial pH of 7.0, temperature of 37°C and rotational speed of 180 rev/min were optimum processing parameters for diacetyl
production. The maximum yield of diacetyl could reach 1.35 g l−1 in a 5-l bioreactor. These results showed an enhancement of the non-enzymatic oxidative decarboxylation of α-acetolactate
and a decrease in the activities of Ald, Ldh and Dr as a consequence of diacetyl accumulation in UV-3. 相似文献
19.
Tributyl phosphate (TBP) is widely used in nuclear fuel processing and other waste generating chemical industries. Although TBP is bacteriostatic, some microbes are resistant to it and may degrade it. Under dark aerobiosis, purple non-sulfur photosynthetic bacteria degraded up to 0.6 mM TBP, initially present at 2 mm, within 3 weeks and under photosynthetic conditions, Rhodopseudomonas palustris degraded 1.6 mM TBP within 3 weeks. The curing of the Rhodopseudomonas palustris endogenous plasmid demonstrated that the genes involved in the TBP degradation are chromosomal. 相似文献
20.
Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes. 相似文献