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1.
Xin-chun Mo Chun-lan Chen Hao Pang Yi Feng Jia-xun Feng 《Applied microbiology and biotechnology》2010,87(6):2137-2146
A metagenomic library containing ca. 3.06 × 108 bp insert DNA was constructed from a rice straw degrading enrichment culture. A xylanase gene, umxyn10A, was cloned by screening the library for xylanase activity. The encoded enzyme Umxyn10A showed 58% identity and 73% similarity
with a xylanase from Thermobifida fusca YX. Sequence analyses showed that Umxyn10A contained a glycosyl hydrolase family 10 catalytic domain. The gene was expressed
in Escherichia coli, and the recombinant enzyme was purified and characterized biochemically. Recombinant Umxyn10A was highly active toward xylan.
However, the purified enzyme could slightly hydrolyze β-1,3/4-glucan and β-1,3/6-glucan. Umxyn10A displayed maximal activity
toward oat spelt xylan at a high temperature (75°C) and weak acidity (pH 6.5). The K
m
and V
max of Umxyn10A toward oat spelt xylan were 3.2 mg ml−1 and 0.22 mmol min−1 mg−1 and were 2.7 mg ml−1 and 1.0 mmol min−1 mg−1 against birchwood xylan, respectively. Metal ions did not appear to be required for the catalytic activity of this enzyme.
The enzyme Umxyn10A could efficiently hydrolyze birchwood xylan to release xylobiose as the major product and a negligible
amount of xylose. The xylanase identified in this work may have potential application in producing xylobiose from xylan. 相似文献
2.
3.
A recombinant putative glycoside hydrolase from Caldicellulosiruptor saccharolyticus was purified with a specific activity of 12 U mg−1 by heat treatment and His-Trap affinity chromatography, and identified as a single 56 kDa band upon SDS-PAGE. The native
enzyme is a dimer with a molecular mass of 112 kDa as determined by gel filtration. The enzyme exhibited its highest activity
when debranched arabinan (1,5-α-l-arabinan) was used as the substrate, demonstrating that the enzyme was an endo-1,5-α-l-arabinanase. The K
m, k
cat, and k
cat/K
m values were 18 mg ml−1, 50 s−1, and a 2.8 mg ml−1 s−1, respectively. Maximum enzyme activity was at pH 6.5 and 75°C. The half-lives of the enzyme at 65, 70 and 75°C were 2440,
254 and 93 h, respectively, indicating that it is the most thermostable of the known endo-1,5-α-l-arabinanases. 相似文献
4.
Caroline Mirande Pascale Mosoni Christel Béra-Maillet Annick Bernalier-Donadille Evelyne Forano 《Applied microbiology and biotechnology》2010,87(6):2097-2105
A xylanase gene xyn10A was isolated from the human gut bacterium Bacteroides xylanisolvens XB1A and the gene product was characterized. Xyn10A is a 40-kDa xylanase composed of a glycoside hydrolase family 10 catalytic
domain with a signal peptide. A recombinant His-tagged Xyn10A was produced in Escherichia coli and purified. It was active on oat spelt and birchwood xylans and on wheat arabinoxylans. It cleaved xylotetraose, xylopentaose,
and xylohexaose but not xylobiose, clearly indicating that Xyn10A is a xylanase. Surprisingly, it showed a low activity against
carboxymethylcellulose but no activity at all against aryl-cellobioside and cellooligosaccharides. The enzyme exhibited K
m and V
max of 1.6 mg ml−1 and 118 μmol min−1 mg−1 on oat spelt xylan, and its optimal temperature and pH for activity were 37°C and pH 6.0, respectively. Its catalytic properties
(k
cat/K
m = 3,300 ml mg−1 min−1) suggested that Xyn10A is one of the most active GH10 xylanase described to date. Phylogenetic analyses showed that Xyn10A
was closely related to other GH10 xylanases from human Bacteroides. The xyn10A gene was expressed in B. xylanisolvens XB1A cultured with glucose, xylose or xylans, and the protein was associated with the cells. Xyn10A is the first family 10
xylanase characterized from B. xylanisolvens XB1A. 相似文献
5.
L Mao P Meng C Zhou L Ma G Zhang Y Ma 《World journal of microbiology & biotechnology》2012,28(3):777-784
A new xylanase gene, named xyn186, was cloned by the genome-walking PCR method from the Alternaria sp. HB186. The sequence of xyn186 contains a 748 bp open reading frame separated by one intron with the size of 52 bp. The cDNA was obtained by DpnI-mediated intron deletion. The cDNA was cloned into pHBM905A and transformed into Pichia pastoris GS115 to screen xylanase-secreting transformants on RBB-xylan plates. The molecular mass of the enzyme was estimated to be
23 kDa on SDS-PAGE. The optimal pH and temperature of the purified enzyme is 6 and 50°C, respectively. The K
m and V
max valued for birchwood xylan are 1.404 mg ml−1 and 0.2748 mmol min−1 mg−1, respectively. The inhibitory effects of various metal ions were investigated, Cu2+ and Hg2+ ions inhibited most of the enzyme activity. The gene copy number of xyn186 in the genome of P. pastoris was estimated as two by the Real-time PCR. To date, xyn186 gene is the first xylanase gene cloned from the genus Alternaria. 相似文献
6.
Basaran Pervin Basaran Nese Hang Yong D. 《World journal of microbiology & biotechnology》2000,16(6):545-550
Pichia stipitis strain NRRL Y-11,543 was mutagenized with N-methyl-N′-nitro-N-nitrosoguanidine (NTG) to improve xylanolytic activity. A total of 20,000 mutants were screened for xylanase overproduction
by observing the clear zones around the colonies on remazol-briliant-blue-xylan (RBB-xylan)-containing agar. Of 94 mutants
isolated 11 of them were found to have enhanced xylanase activity compared to the parental strain. The most active mutant
NP54376 had superior properties to the wild type which included: double the enzyme activity of wild type, a shorter generation
time of 2.22 h compared to 3.13 h when grown on xylan, and an enhanced growth and yield of xylanase when low levels of xylose
were added to the medium. Zymogram analysis of the crude enzyme preparations from both NP54376 and the wild type by isoelectric
focusing showed multiple bands ranging between pI 4.2 and 7.4. No significant difference was observed in the K
m and V
max values of the parental strain and NP54376. K
m and V
max values of xylanase for birchwood xylan were 4.2 mg ml−1 and 0.08 μmol min−1 mg−1 of protein, respectively.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
7.
He Jun Yu Bing Zhang Keying Ding Xuemei Chen Daiwen 《Indian journal of microbiology》2009,49(2):188-195
The strain of Trichoderma reesei Rut C-30 was subjected to mutation after treatment with N-methyl-N′-nitro-N-nitrosoguanidine (NG) for 6 h followed by UV irradiation for 15 min. Successive mutants showed enhanced cellulase production,
clear hydrolysis zone and rapid growth on Avicel-containing plate. Particularly, the mutant NU-6 showed approximately two-fold
increases in activity of both FPA and CMCase in shake flask culture when grown on basal medium containing peptone (1%) and
wheat bran (1%). The enzyme production was further optimized using eight different media. When a mixture of lactose and yeast
cream was used as cellulase inducer, the mutant NU-6 yielded the highest enzyme and cell production with a FPase activity
of 6.2 U ml−1, a CMCase activity of 54.2 U ml−1, a β-glucosidase activity of 0.39 U ml−1, and a fungal biomass of 12.6 mg ml−1. It deserved noting that the mutant NU-6 also secreted large amounts of xylanases (291.3 U ml−1). These results suggested that NU-6 should be an attractive producer for both cellulose and xylanase production. 相似文献
8.
The effect of agitation and aeration on the growth and antibiotic production by Xenorhabdus nematophila YL001 grown in batch cultures were investigated. Efficiency of aeration and agitation was evaluated through the oxygen mass
transfer coefficient (K
L
a). With increase in K
L
a, the biomass and antibiotic activity increased. Activity units of antibiotic and dry cell weight were increased to 232 U ml−1 and 19.58 g l−1, respectively, productivity in cell and antibiotic was up more than 30% when K
L
a increased from 115.9 h−1 to 185.7 h−1. During the exponential growth phase, DO concentration was zero, the oxygen supply was not sufficient. So, based on process
analysis, a three-stage oxygen supply control strategy was used to improved the DO concentration above 30% by controlling
the agitation speed and aeration rate. The dry cell weight and activity units of antibiotic were further increased to 24.22 g l−1 and 249 U ml−1, and were improved by 24.0% and 7.0%, compared with fermentation at a constant agitation speed and a constant aeration rate
(300 rev min−1, 2.5 l min−1). 相似文献
9.
Vikramathithan J Muthuraman P Ravikumar S Shayamala S Kumar GN Srikumar K 《The protein journal》2012,31(2):141-149
Two thermostable xylanase isoforms T60 and T80 were purified to homogeneity from the cladodes of the xerophytic Cereus pterogonus plant species. After three consecutive purification steps, the specific activity of T60 and T80 isoforms were found to be 178.6 and 216.2 U mg−1 respectively. The molecular mass of both isoforms was determined to be 80 kDa. The optimum temperature for T60 and T80 xylanase isoforms were 60 and 80 °C respectively. The pH was 5.0 for both isoforms. The presence of divalent metal ions (10 mM
Co2+) showed stimulatory effects of both catalytic activities, where as in the presence of Hg2+, Cd2+, Cu2+ showed inhibitory effect on these activities at all concentrations studied. The thermodynamic analysis of xylanase activity
using denaturation kinetics and the presence divalent cations at 30–100 °C, showed lower ΔH, ΔS, and ΔG values at all the temperatures investigated. The melting temperature of purified T80 xylanase isoform as determined by TG/DTA analysis and it showed the unfolding temperature was 80 °C. The g value and hyperfine
(A) value purified xylanase T80 isoform was 2.017 and 10.80 respectively. Immunoblot analysis with antiserum raised against the purified T80 xylanase isoforms revealed single immunolgically related polypeptides of 80 kDa, identical with the polypeptide band produced
on SDS-PAGE. The results of double immunodiffusion against the T80 isoforms showed a single precipitin line indicating that the serum used was specific to these xylanase isoforms. The kinetic
and thermodynamic properties suggested that xylanase from C. pterogonus may have a potential usage in various industries. 相似文献
10.
Kim HT Ko HJ Kim N Kim D Lee D Choi IG Woo HC Kim MD Kim KH 《Biotechnology letters》2012,34(6):1087-1092
A gene, alg7D, from Saccharophagus degradans, coding for a putative alginate lyase belonging to the family of polysaccharide lyase-7, was overexpressed in Escherichia coli. The properties of the recombinant Alg7D were characterized. The enzyme endolytically depolymerized alginate by β-elimination
into oligo-alginates with degrees of polymerization of 2–5. Its activity was maximal at 50°C and pH 7 and was slightly increased
in the presence of Na+. The K
M
, V
max
, k
cat
, and k
cat
/K
M
values were: 3 mg ml−1, 6.2 U mg−1, 1.9 × 10−2 s−1, and 6.3 × 10−3 mg−1 ml s−1, respectively. 相似文献
11.
Hyaluronic acid production by recombinant <Emphasis Type="Italic">Lactococcus lactis</Emphasis> 总被引:1,自引:0,他引:1
A gene encoding a new xylanase, named xynZG, was cloned by the genome-walking PCR method from the nematophagous fungus Plectosphaerella cucumerina. The genomic DNA sequence of xynZG contains a 780 bp open reading frame separated by two introns with the sizes of 50 and 46 bp. To our knowledge, this would
be the first functional gene cloned from P. cucumerina. The 684 bp cDNA was cloned into vector pHBM905B and transformed into Pichia pastoris GS115 to select xylanase-secreting transformants on RBB-xylan containing plate. The optimal secreting time was 3 days at
25°C and enzymatic activities in the culture supernatants reached the maximum level of 362 U ml−1. The molecular mass of the enzyme was estimated to be 19 kDa on SDS-PAGE. The optimal pH and temperature of the purified
enzyme is 6 and 40°C, respectively. The purified enzyme is stable at room temperature for at least 10 h. The K
m and V
max values for birchwood xylan are 2.06 mg ml−1 and 0.49 mmol min−1mg−1, respectively. The inhibitory effects of various mental ions were investigated. It is interesting to note that Cu2+ ion, which strongly inhibits most other xylanases studied, reduces enzyme activity by only 40%. Furthermore, enzyme activity
is unaffected by EDTA even at a concentration of 5 mM. 相似文献
12.
Behzad Ahmadi Khoshnood Alizadeh Jaime A. Teixeira da Silva 《Plant Cell, Tissue and Organ Culture》2012,109(3):525-533
The effects of three periods of incubation (10, 20 and 30 min) at different levels of bleomycin (0, 0.1, 0.2, 0.3, 0.4 and
0.5 μg ml−1), as well as three periods of exposure (12, 24 and 48 h) to different levels of the anti-auxin p-chlorophenoxyisobutyric acid (PCIB), including 1, 2, 3, 4 and 5 mg l−1, on microspore embryogenesis of rapeseed cv. ‘Amica’ were investigated. Microspore embryogenesis was significantly enhanced
following 20 min treatment with 0.2 μg ml−1 bleomycin compared with untreated cultures. Highest embryo yield (163 embryos Petri dish−1) was observed with 24 h treatment of 4 mg l−1 PCIB. The highest percentage of secondary embryogenesis was observed on B5 medium containing 0.15 mg l−1 of gibberellic acid (GA3) and 0.2 mg l−1 6-benzyladenine (BA) in 4–6 mm microspore-derived embryos (MDEs). Most callus formed on B5 medium containing 0.15 mg l−1 GA3, 0.1 mg l−1 BA and 0.1 mg l−1 indole-3-acetic acid (IAA) when 4–6 mm embryos were used. Regeneration was highest on B5 medium containing 0.05 mg l−1 GA3 or 0.1 mg l−1 BA and 0.2 mg l−1 IAA with 2–4 mm embryos. Microspore embryogenesis and plant regeneration could be improved by both bleomycin and PCIB when
the appropriate MDE length and phytohormone level were selected. 相似文献
13.
Jeyagowri Kiddinamoorthy Alfredo J. Anceno Gulelat D. Haki Sudip K. Rakshit 《World journal of microbiology & biotechnology》2008,24(5):605-612
Production of extracellular xylanase from Bacillus sp. GRE7 using a bench-top bioreactor and solid-state fermentation (SSF) was attempted. SSF using wheat bran as substrate
and submerged cultivation using oat-spelt xylan as substrate resulted in an enzyme productivity of 3,950 IU g−1 bran and 180 IU ml−1, respectively. The purified enzyme had an apparent molecular weight of 42 kDa and showed optimum activity at 70°C and pH
7. The enzyme was stable at 60–80°C at pH 7 and pH 5–11 at 37°C. Metal ions Mn2+ and Co2+ increased activity by twofold, while Cu2+ and Fe2+ reduced activity by fivefold as compared to the control. At 60°C and pH 6, the K
m for oat-spelt xylan was 2.23 mg ml−1 and V
max was 296.8 IU mg−1 protein. In the enzymatic prebleaching of eucalyptus Kraft pulp, the release of chromophores, formation of reducing sugars
and brightness was higher while the Kappa number was lower than the control with increased enzyme dosage at 30% reduction
of the original chlorine dioxide usage. The thermostability, alkali-tolerance, negligible presence of cellulolytic activity,
ability to improve brightness and capacity to reduce chlorine dioxide usage demonstrates the high potential of the enzyme
for application in the biobleaching of Kraft pulp. 相似文献
14.
M.I. Rajoka Khalil-ur- Rehman Tehmina Tabish M.A. Zia 《World journal of microbiology & biotechnology》2006,22(3):289-291
Purified uricase from a caprine kidney, possessed K
m
and V
max values of 1.1 mg ml−1 and 3512 IU (mg protein)−1 for uric acid hydrolysis, respectively. The optimum temperature and pH for catalytic activity were 40 °C and 8.5, respectively.
The activation energy for formation of ES complex was 13.6 kJ mol−1. Enthalpy (ΔH*), entropy of activation (ΔS*) and Gibbs free energy demand of uricase inactivation were 62.8 kJ mol−1, −102 J mol−1 K−1 and 104.3 kJ mol−1, respectively. Gibbs free enrgy demand for substrate binding and transition state stabilization were also determined which
were comparable with those for themostable enzymes. 相似文献
15.
Teodora Janković Branka Vinterhalter Dijana Krstić-Milošević Radomirka Nikolić Dragan Vinterhalter Slobodan Milosavljević 《Acta Physiologiae Plantarum》2011,33(4):1515-1520
Shoot cultures of Gentianella bulgarica established from seedling epicotyls were maintained on MS medium supplemented with BA 0.2 mg l−1 + NAA 0.1 mg l−1. Cultures were prone to precocious flowering requiring the use of small shoot buds for multiplication purposes. The contents
of three xanthone compounds identified as DGL, BGL, and DMB, in different plant material were determined by HPLC. The analysis
revealed that the production of xanthones was affected by different concentrations of BA in medium. Shoot cultures grown at
higher BA concentrations contained more DGL than material grown in nature. The concentrations of other two xanthones were
lower in shoot cultures than in plants from nature. The radical scavenging activity of plant extracts and xanthones was investigated
by DPPH test. Samples from plants grown in nature showed the highest activity (IC50 = 0.26 mg ml−1), while the extracts of shoot cultures grown in media with higher concentrations of BA showed moderate activities (IC50 from 1.6 to 4.4 mg ml−1). 相似文献
16.
The interaction between benzophenone (BP) and bovine serum albumin (BSA) was investigated by the methods of fluorescence spectroscopy
combined with UV–Vis absorption and circular dichroism (CD) measurements under simulative physiological conditions. The experiment
results showed that the fluorescence quenching of BSA by BP was resulted from the formation of a BP–BSA complex and the corresponding
association constants (K
a) between BP and BSA at four different temperatures had been determined using the modified Stern–Volmer equation. The enthalpy
change (ΔH) and entropy change (ΔS) were calculated to be –43.73 kJ mol−1 and −53.05 J mol−1 K−1, respectively, which suggested that hydrogen bond and van der Waals force played major roles in stabilizing the BP–BSA complex.
Site marker competitive experiments indicated that the binding of BP to BSA primarily took place in site I (sub-domain IIA).
The conformational investigation showed that the presence of BP decreased the α-helical content of BSA and induced the slight unfolding of the polypeptides of protein, which confirmed some micro-environmental
and conformational changes of BSA molecules. 相似文献
17.
Huiying Luo Jun Yang Jiang Li Pengjun Shi Huoqing Huang Yingguo Bai Yunliu Fan Bin Yao 《Applied microbiology and biotechnology》2010,86(6):1829-1839
We cloned and sequenced a xylanase gene named xylD from the acidophilic fungus Bispora sp. MEY-1 and expressed the gene in Pichia pastoris. The 1,422-bp full-length complementary DNA fragment encoded a 457-amino acid xylanase with a calculated molecular mass of
49.8 kDa. The mature protein of XYLD showed high sequence similarity to both glycosyl hydrolase (GH) families 5 and 30 but
was more homologous to members of GH 30 based on phylogenetic analysis. XYLD shared the highest identity (49.9%) with a putative
endo-1,6-β-d-glucanase from Talaromyces stipitatus and exhibited 21.1% identity and 34.3% similarity to the well-characterized GH family 5 xylanase from Erwinia chrysanthemi. Purified recombinant XYLD showed maximal activity at pH 3.0 and 60 °C, maintained more than 60% of maximal activity when
assayed at pH 1.5–4.0, and had good thermal stability at 60 °C and remained stable at pH 1.0–6.0. The enzyme activity was
enhanced in the presence of Ni2+ and β-mercaptoethanol and inhibited by some metal irons (Hg2+, Cu2+, Pb2+, Mn2+, Li+, and Fe3+) and sodium dodecyl sulfate. The specific activity of XYLD for beechwood xylan, birchwood xylan, 4-O-methyl-d-glucuronoxylan, and oat spelt xylan was 2,463, 2,144, 2,020, and 1,429 U mg−1, respectively. The apparent K
m and V
max values for beechwood xylan were 5.6 mg ml−1 and 3,622 μmol min−1 mg−1, respectively. The hydrolysis products of different xylans were mainly xylose and xylobiose. 相似文献
18.
Naeem Ali Abdul Hameed Safia Ahmed Abdul G. Khan 《World journal of microbiology & biotechnology》2008,24(7):1067-1072
The fungal strain, Aspergillus niger SA1, isolated from textile wastewater sludge was screened for its decolorization ability for four different textile dyes.
It was initially adapted to higher concentration of dyes (10–1,000 mg l−1) on solid culture medium after repeated sub-culturing. Maximum resistant level (mg l−1) sustained by fungal strain against four dyes was in order of; Acid red 151 (850) > Orange II (650) > Drimarene blue K2RL (550) > Sulfur black (500). The apparent dye removal for dyes was seen largely due to biosorption/bioadsorption into/onto
the fungal biomass. Decolorization of Acid red 151, Orange II, Sulfur black and Drimarine blue K2RL was 68.64 and 66.72, 43.23 and 44.52, 21.74 and 28.18, 39.45 and 9.33% in two different liquid media under static condition,
whereas, it was 67.26, 78.08, 45.83 and 13.74% with 1.40, 1.73, 5.16 and 1.87 mg l−1 of biomass production under shaking conditions respectively in 8 days. The residual amount (mg l−1) of the three products (α-naphthol, sulfanilic acid and aniline) kept quite low i.e., ≤2 in case AR 151 and Or II under shaking
conditions. Results clearly elucidated the role of Aspergillus niger SA1 in decolorizing/degrading structurally different dyes into basic constituents. 相似文献
19.
Cui Fengjie Li Yin Liu Zhiqiang Zhao Hui Ping Lifeng Ping Liying Yang Yinan Xue Yaping Yan Lijiao 《World journal of microbiology & biotechnology》2009,25(4):721-725
The objective of this study was to maximize production of xylanase by a newly isolated strain Penicillium thiersii ZH-19. Response surface methodology was employed to study the effects of significant factors such as pH, temperature, xylan
concentration, and cultivation time, on the production of xylanase by Penicillium thiersii ZH-19. The optimal fermentation parameters for enhanced xylanase production were found to be pH 7.72, temperature 24.8°C, xylan
13.2 g l−1 and the fermentation time 125.8 h. The model predicted a xylanase activity of 75.24 U ml−1. Verification of the optimization showed that the maximum xylanase production reached 73.50 U mL−1 in the flask experiments and 80.23 U mL−1 in the scale of 15-L fermenter under the optimal condition. 相似文献
20.
In recent years, the biotechnological use of xylanases has grown remarkably. To efficiently produce xylanase for food processing
and other industry, a codon-optimized recombinant xylanase gene from Streptomyces sp. S38 was synthesized and extracellularly expressed in Pichia pastoris under the control of AOX1 promoter. SDS-PAGE and activity assay demonstrated that the molecular mass of the recombinant xylanase was estimated to be
25 kDa, the optimum pH and optimum temperature were 5.5 and 50°C, respectively. In shake flask culture, the specific activity
of the xylanase activity was 5098.28 U/mg. The K
m
and V
max
values of recombinant xylanase were 11.0 mg/ml and 10000 μmol min−1 mg−1, respectively. In the presence of metal ions such as Ca2+, Cu2+, Cr3+ and K+, the activity of the enzyme increased. However, strong inhibition of the enzyme activity was observed in the presence of
Hg2+. This is the first report on the expression properties of a recombinant xylanase gene from the Streptomyces sp. S38 using Pichia pastoris. The attractive biochemical properties of the recombinant xylanase suggest that it may be a useful candidate for variety
of commercial applications. 相似文献