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1.
Thermolabile xylanase of the Antarctic yeast Cryptococcus adeliae: production and properties 总被引:4,自引:0,他引:4
Xylanase production by the Antarctic psychrophilic yeast Cryptococcus adeliae was increased 4.3 fold by optimizing the culture medium composition using statistical designs. The optimized medium containing
24.2 g l−1 xylan and 10.2 g l−1 yeast extract and having an initial pH of 7.5 yielded xylanase activity at 400 nkat (nanokatal) ml−1 after 168-h shake culture at 4°C. In addition, very little endoglucanase, β-mannanase, β-xylosidase, β-glucosidase, α-l-arabinofuranosidase, and no filter paper cellulase activities were detected. Among 12 carbon sources tested, maximum xylanase
activity was induced by xylan, followed by lignocelluloses such as steamed wheat straw and alkali-treated bagasse. The level
of enzyme activity produced on other carbon sources appeared to be constitutive. Among the complex organic nitrogen sources
tested, the xylanase activity was most enhanced by yeast extract, followed by soymeal, Pharmamedia (cotton seed protein),
and Alburex (potato protein). A batch culture at 10°C in a 5-l fermenter (3.5-1 working volume) using the optimized medium
gave 385 nkat at 111 h of cultivation. The crude xylanase showed optimal activity at pH 5.0–5.5 and good stability at pH 4–9
(21 h at 4°C). Although the enzyme was maximally active at 45°–50°C, it appeared very thermolabile, showing a half-life of
78 min at 35°C. At 40°–50°C, it lost 71%–95% activity within 5 min. This is the first report on the production as well as
on the properties of thermolabile xylanase produced by an Antarctic yeast.
Received: December 10, 1999 / Accepted: March 23, 2000 相似文献
2.
Penicillium funiculosum NRRL 13033 produced β-glucosidase and β-xylosidase activities when grown on wheat straw. The addition of some inducers (individually or in combination) to the fermentation medium were tested for the production of both enzymes. The relation of mycelial bound enzyme to cell free enzyme was studied during incubation period of fermentation. The optimum activity of β-glucosidase and β-xylosidase were found to be in the pH 4.5 using phosphate-citrate buffer at 50°C for 60 min and at 55°C for 40 min respectively. β-Glucosidase lost about 40% of its original activity by heating to 65°C for 60 min, while, β-xylosidase activity was found to be nearly stable with the same treatment. Both enzyme activities were greatly inhibited when 1.0% (w/v) of xylose and glucose were added to the assay mixture. 相似文献
3.
Twenty Aspergillus strains were evaluated for production of extracellular cellulolytic and xylanolytic activities. Aspergillus brasiliensis, A. niger and A. japonicus produced the highest xylanase activities with the A. brasiliensis and A. niger strains producing thermostable β-xylosidases. The β-xylosidase activities of the A. brasiliensis and A. niger strains had similar temperature and pH optima at 75°C and pH 5 and retained 62% and 99%, respectively, of these activities
over 1 h at 60°C. At 75°C, these values were 38 and 44%, respectively. Whereas A. niger is a well known enzyme producer, this is the first report of xylanase and thermostable β-xylosidase production from the newly
identified, non-ochratoxin-producing species A. brasiliensis. 相似文献
4.
The anaerobic fungus Anaeromyces mucronatus KF8 grown in batch culture on M10 medium with rumen fluid and microcrystalline cellulose as carbon source produced a broad
range of enzymes requisite for degradation of plant structural and storage saccharides including cellulase, endoglucanase,
xylanase, α-xylosidase, β-xylosidase, α-glucosidase, β-glucosidase, β-galactosidase, mannosidase, cellobiohydrolase, amylase,
laminarinase, pectinase and pectate lyase. These enzymes were detected in both the intra- and extracellular fractions, but
production into the medium was prevalent with the exception of intracellular β-xylosidase, chitinases, N-acetylglucosaminidase, and lipase. Xylanase activity was predominant among the polysaccharide hydrolases. Extracellular production
of xylanase was stimulated by the presence of cellobiose and oat spelt xylan. Zymogram of xylanases of strain KF8 grown on
different carbon sources revealed several isoforms of xylanases with approximate molar masses ranging from 26 to 130 kDa. 相似文献
5.
Purification and partial characterization οf a new β-xylosidase from
Humicola grisea var. thermoidea
T. Iembo M.O. Azevedo C. Bloch Jr. E.X.F. Filho 《World journal of microbiology & biotechnology》2006,22(5):475-479
Summary The thermophilic fungus Humicola grisea var. thermoidea produces a mycelium-associated β-xylosidase activity when grown in liquid-state cultures on media containing oat spelt xylan
as the carbon source. The β-xylosidase was purified to apparent homogeneity by gel filtration and anion exchange chromatography.
Its molecular weight was 37 and 50 kDa, as determined by MALDI/TOF mass spectrometry and SDS-PAGE, respectively. The purified
enzyme exhibited maximum activity at 55 °C and pH 6.5. It was also active at pH 8.8, retaining 60% of its activity after 6 h
of incubation at 50 °C. β-xylosidase was strongly inactivated by NBS and slightly activated by DTT and β-mercaptoethanol.
The enzyme was highly specific for PNPX as the substrate. The purified β-xylosidase showed K
m and V
max values of 1.37 mM and 12.98 IU ml−1, respectively. 相似文献
6.
Chang L Ding M Bao L Chen Y Zhou J Lu H 《Applied microbiology and biotechnology》2011,90(6):1933-1942
A new gene, RuCelA, encoding a bifunctional xylanase/endoglucanase, was cloned from a metagenomic library of yak rumen microorganisms. RuCelA
showed activity against xylan and carboxymethylcellulose (CMC), suggesting bifunctional xylanase/endoglucanase activity. The
optimal conditions for xylanase and endoglucanase activities were 65°C, pH 7.0 and 50°C, pH 5.0, respectively. In addition,
the presence of Co+ and Co2+ can greatly improve RuCelA's endoglucanase activity, while inhibits its xylanase activity. Further examination of substrate
preference showed a higher activity against barley glucan and lichenin than against xylan and CMC. Using xylan and barley
glucan as substrates, RuCelA displayed obvious synergistic effects with β-1,4-xylosidase and β-1,4-glucosidase. Generation
of soluble oligosaccharides from lignocellulose is the key step in bioethanol production, and it is greatly notable that RuCelA
can produce xylo-oligosaccharides and cello-oligosaccharides in the continuous saccharification of pretreated rice straw,
which can be further degraded into fermentable sugars. Therefore, the bifunctional RuCelA distinguishes itself as an ideal
candidate for industrial applications. 相似文献
7.
M. Michelin S. C. Peixoto-Nogueira J. H. A. Betini T. M. da Silva J. A. Jorge H. F. Terenzi M. L. T. M. Polizeli 《Bioprocess and biosystems engineering》2010,33(7):813-821
Aspergillus terricola and Aspergillus ochraceus, isolated from Brazilian soil, were cultivated in Vogel and Adams media supplemented with 20 different carbon sources, at
30 °C, under static conditions, for 120 and 144 h, respectively. High levels of cellulase-free xylanase were produced in birchwood
or oat spelt xylan-media. Wheat bran was the most favorable agricultural residue for xylanase production. Maximum activity
was obtained at 60 °C and pH 6.5 for A. terricola, and 65 °C and pH 5.0 for A. ochraceus. A. terricola xylanase was stable for 1 h at 60 °C and retained 50% activity after 80 min, while A. ochraceus xylanase presented a t
50 of 10 min. The xylanases were stable in an alkali pH range. Biobleaching of 10 U/g dry cellulose pulp resulted in 14.3% delignification
(A. terricola) and 36.4% (A. ochraceus). The brightness was 2.4–3.4% ISO higher than the control. Analysis in SEM showed defibrillation of the microfibrils. Arabinase
traces and β-xylosidase were detected which might act synergistically with xylanase. 相似文献
8.
The culture-medium composition was optimised, on a shake-flask scale, for simultaneous production of high activities of endoglucanase
and β-glucosidase by Thermoascus aurantiacus using statistical factorial designs. The optimised medium containing 40.2 g l−1 Solka Floc as the carbon source and 9 g l−1 soymeal as the organic nitrogen source yielded 1130 nkat ml−1 endoglucanase and 116 nkat ml−1β-glucosidase activities after 264 h as shake cultures. In addition, good levels of β-xylanase (3479 nkat ml−1) and low levels of filter-paper cellulase, β-xylosidase, α-l-arabinofuranosidase, β-mannanase, β-mannosidase, α-galactosidase and β-galactosidase were detected. Batch fermentation in
a 5-l laboratory fermentor using the optimised medium allowed the production of 940 nkat ml−1 endoglucanase and 102 nkat ml−1β-glucosidase in 192 h. Endoglucanase and β-glucosidase showed optimum activity at pH 4.5 and pH 5, respectively, and they
displayed optimum activity at 75 °C. Endoglucanase and β-glucosidase showed good stability at pH values 4–8 and 4–7, respectively,
after a prolonged incubation (48 h at 50 °C). Endoglucanase had half-lives of 98 h at 70 °C and 4.1 h at 75 °C, while β-glucosidase
had half-lives of 23.5 h at 70 °C and 1.7 h at 75 °C. Alkali-treated bagasse, steam-treated wheat straw, Solka floc and Sigmacell
50 were 66, 48.5, 33.5 and 14.4% hydrolysed by a crude enzyme complex of T. aurantiacus in 50 h.
Received: 12 November 1999 / Accepted: 14 November 1999 相似文献
9.
A. I. El-Diwany M. H. Selim A. M. El-Shafei 《Applied microbiology and biotechnology》1987,26(6):552-554
Summary β-Glucosidase production by Debaryomyces vanrigii and Debaryomyces hansenii was studied using two media. Cellobiose was found to stimulate the biosynthesis of the enzyme, while NH4NO3 (1.0 g/l) and NH4Cl (1.26 g/l) were the best nitrogen sources for D. hansenii and D. vanrigii respectively. Optimal conditions for enzyme activity were established in relation to pH, temperature and enzyme stability.
Thermal and pH stability studies show that β-glucosidase from D. vanrigii was more stable at pH 4.5–5.0 at 50°C, while that enzyme from D. hansenii was stable at pH 6.5 at 35°C. This feature may be advantageous in the commercial application by hydrolysing cellobiose, the
potent inhibitor of cellulose solubilizing enzymes. 相似文献
10.
β-Glucosidase from almonds (EC 3.2.1.21) was covalently immobilized by a two-step technique. In the first step, double bonds
were introduced into the β-glucosidase by derivatization with itaconic anhydride. In separate studies with α-N-protected l-amino acids, it was established that itaconic anhydride acylated mainly primary amino groups of lysines and, to a much lesser
extent hydroxyl groups of tyrosines and sulfhydryl groups of cysteines. The acylated β-glucosidase showed no loss of activity
and the K
m decreased from 3.6 mM to 2.6 mM when p-nitrophenyl β-d-glucopyranoside was used as the substrate. In the second step, the derivatized β-glucosidase was co-polymerized radically
with N,N′-methylenebisacrylamide in buffer solution. The resulting acrylamide immobilizate possessed a much better storage stability
at 30–56 °C when compared to β-glucosidase immobilized on Eupergit C. However, the specific activity was higher with the Eupergit
immobilizate. Free and acrylamide-immobilized β-glucosidase were used for glucosylation of chloramphenicol by transglucosylation
in 20% (v/v) acetonitrile at 37 °C. The acrylamide immobilizate demonstrated a great enhancement of stability and approximately
50% more chloramphenicol β-glucoside was obtained after 5 h.
Received: 22 September 1997 / Accepted: 28 October 1997 相似文献
11.
Yongling Qin Haiyan He Nan Li Min Ling Zhiqun Liang 《World journal of microbiology & biotechnology》2010,26(11):1991-1997
A thermostable cellulase-producing fungus, HML 0278, was identified as Fusarium chlamydosporum by morphological characteristics and ITS rDNA sequence analysis. HML 0278 produced extracellular cellulases in solid-state
fermentation using sugar cane bassage as the carbon source. Native-PAGE analysis demonstrated that this fungus strain was
capable of producing the three major components of cellulases and xylanase, with a yield of 281.8 IU/g for CMCase, 182.4 IU/g
for cellobiohydrolase, 135.2 IU/g for β-glucosidase, 95.2 IU/g for filter paper activity, and 4,720 IU/g for xylanase. More
importantly, the CMCase and β-glucosidase produced by HML 0278 showed stable enzymatic activities within pH 4–9 and pH 4–10,
and at temperatures below 70 and 60°C, respectively. 相似文献
12.
The influence of different cultivation conditions on β-glucosidase production and of some parameters on the activity and stability
of this enzyme were studied inNectria catalinensis. Maximal β-glucosidase production was achieved with ammonium nitrate (0.5 g N/L) as nitrogen source. Tween 80, Tween 20 and
Triton X-100 increased β-glucosidase yields, Tween 80 was the most effective for enzyme release and growth at a concentration
of 3.4 mmol/L. On the other hand, Tween 20 and Triton X-100 had an inhibitory effect onN. catalinensis growth. A temperature of 23°C and an initial pH of cultures of 6.5 were optimal for biomass and β-glucosidase production.
Under optimal cultural conditions (ammonium nitrate, 0.5 g N/L; Tween 80, 3.4 mmol/L; 23°C; initial pH 6.5) the β-glucosidase
yield was increased more than five fold respect to the initial state. Optimal temperature for β-glucosidase activity was 45°C,
the initial activity dropped 60 % after 6 h of incubation at this temperature. Optimal pH for enzyme activity was 5.3. At
this pH the β-glucosidase was completely stable after 3 d of incubation. TheV andK
m values calculated from Lineweaver-Burk and Eadie-Hofstee plots were 0.23 μmol 4-nitrophenol per min per mg of protein and
0.25 mmol 4-nitrophenol β-d-glucopyranoside per L, respectively. The activation energy according to Arrhenius plot was 49.6 KJ/mol. 相似文献
13.
Hong SY Lee JS Cho KM Math RK Kim YH Hong SJ Cho YU Cho SJ Kim H Yun HD 《Biotechnology letters》2007,29(6):931-936
An artificial bifunctional enzyme, cellulase-β-glucosidase, was prepared by gene fusion from the hyperthermophilic bacterium
Thermotoga maritima MSB8. The fusion protein exhibited both cellulase (Cel5C) and β-glucosidase (BglB) activity when the bglB gene was fused to downstream of cel5C, but not when cel5C was fused to downstream of bglB. The specific activity of the bifunctional enzyme was 70% lower than that of cellulase or β-glucosidase. The fusion enzyme
was purified, and the MW was estimated as 114 kDa. The fusion enzyme displayed optimum cellulase activity at pH 8.0 and 70°C
over 30 min, and optimal β-glucosidase activity at pH 7.0 and 80°C over 30 min. 相似文献
14.
A bacterium (strain HC1) capable of assimilating rice bran hemicellulose was isolated from a soil and identified as belonging
to the genus Paenibacillus through taxonomical and 16S rDNA sequence analysis. Strain HC1 cells grown on rice bran hemicellulose as a sole carbon source
inducibly produced extracellular xylanase and intracellular glycosidases such as β-d-glucosidase and β-d-arabinosidase. One of them, β-d-glucosidase was further analyzed. A genomic DNA library of the bacterium was constructed in Escherichia coli and gene coding for β-d-glucosidase was cloned by screening for β-d-glucoside-degrading phenotype in E. coli cells. Nucleotide sequence determination indicated that the gene for the enzyme contained an open reading frame consisting
of 1,347 bp coding for a polypeptide with a molecular mass of 51.4 kDa. The polypeptide exhibits significant homology with
other bacterial β-d-glucosidases and belongs to glycoside hydrolase family 1. β-d-Glucosidase purified from E. coli cells was a monomeric enzyme with a molecular mass of 50 kDa most active at around pH 7.0 and 37°C. Strain HC1 glycosidases
responsible for degradation of rice bran hemicellulose are expected to be useful for structurally determining and molecularly
modifying rice bran hemicellulose and its derivatives. 相似文献
15.
β-Glucosidase is frequently used to supplement cellulase preparations for hydrolysis of cellulosic and lignocellulosic substrates
in order to accelerate the conversion of cellobiose to glucose. Typically, commercial cellulase preparations are deficient
in this enzyme and accumulation of cellobiose leads to product inhibition. This study evaluates the potential for recycling
β-glucosidase by immobilization on a methacrylamide polymer carrier, Eupergit C. The immobilized β-glucosidase had improved
stability at 65 °C, relative to the free enzyme, while the profile of activity versus pH was unchanged. Immobilization resulted
in an increase in the apparent Km from 1.1 to 11 mm and an increase in Vmax from 296 to 2430 μmol mg−1 min−1. The effect of immobilized β-glucosidase on the hydrolysis of cellulosic and lignocellulosic substrates was comparable to
that of the free enzyme when used at the same level of protein. Operational stability of the immobilized β-glucosidase was
demonstrated during six rounds of lignocellulose hydrolysis.
Received 22 August 2005; Revisions requested 20 September 2005; Revisions received 8 November 2005; Accepted 10 November 2005 相似文献
16.
M. V. Semenova M. I. Drachevskaya O. A. Sinitsyna A. V. Gusakov A. P. Sinitsyn 《Biochemistry. Biokhimii?a》2009,74(9):1002-1008
Homogeneous β-xylosidases with molecular mass values 120 and 80 kDa (as shown by SDS-PAGE), belonging to the third family
of glycosyl hydrolases, were isolated by anion-exchange, hydrophobic, and gel-penetrating chromatography from enzyme preparations
based on the fungi Aspergillus japonicus and Trichoderma reesei, respectively. The enzymes exhibit maximal activity in acidic media (pH 3.5–4.0), and temperature activity optimum was 70°C
for the β-xylosidase of A. japonicus and 60°C for the β-xylosidase of T. reesei. Kinetic parameters of p-nitrophenyl β-xylopyranoside and xylooligosaccharide hydrolysis by the purified enzymes were determined, which showed that
β-xylosidase of A. japonicus was more specific towards low molecular weight substrates, while β-xylosidase of T. reesei preferred high molecular weight substrates. The competitive type of inhibition by reaction product (xylose) was found for
both enzymes. The interaction of the enzymes of different specificity upon hydrolysis of glucurono- and arabinoxylans was
found. The β-xylosidases exhibit synergism with endoxylanase upon hydrolysis of glucuronoxylan as well as with α-L-arabinofuranosidase
and endoxylanase upon hydrolysis of arabinoxylan. Addition of β-xylosidases increased efficiency of hydrolysis of plant raw
materials with high hemicellulose content (maize cobs) by the enzymic preparation Celloviridine G20x depleted of its own β-xylosidase. 相似文献
17.
Hydrolytic enzyme(s) production in Micrococcus roseus growing on different cellulosic substrates 总被引:1,自引:0,他引:1
Micrococcus roseus (G12) isolated from higher termite Odontotermes obesus gut exhibited cellulose digesting properties. A lignocellulosic substrate, rice husk induced endoglucanase, β-glucosidase, β-xylanase and β-xylosidase production. Besides rice husk, CMC also induced endoglucanase production. β-Glucosidase activity was quite pronounced when rice husk was supplemented with CMC or cellobiose. Both β-xylanase and β-xylosidase activities could be induced by xylan as well as xylobiose, whereas CMC induced partial activity. Endoglucanase and β-xylanase enzymes were secreted into the culture medium, whereas β-glucosidase and β-xylosidase activities were intracellular in nature. Enzyme production was subject to end product inhibition. The extracellular enzyme(s) possessed the potential to saccharify rice husk, xylan and CMC to reducing sugars. 相似文献
18.
Vichien Kitpreechavanich Mitsunori Hayashi Hidehiko Kumagai Shiro Nagai 《Bioscience, biotechnology, and biochemistry》2013,77(7):1703-1711
A β-xylosidase (β-d-xyloside xylohydrolase, EC 3.2.1.37) and β-glucosidase (β-d-glucoside glucohydrolase, EC 3.2.1.21) extracted from a wheat bran culture of Aspergillus fumigatus were purified up to 90-fold and 131-fold, respectively, by ammonium sulfate precipitation, gel filtration, ion exchange chromatography, and hydroxylapatite chromatography. Molecular weights of the β-xylosidase and β-glucosidase were 360,000 and 380,000, respectively, each consisting of four identical subunits. The isoelectric points of β-xylosidase and β-glucosidase were at pH 5.4 and 4.5, respectively. The optimum temperature for the β-xylosidase was 75°C, being stable up to 65°C for 20 min and for the β-glucosidase was 65°C, being stable up to 60°C for 20 min. The optimum pH for both enzymes was about 4.5, being stable between 2 and 8 at 50°C for 20 min. Both enzymes were inhibited by Fe3+, Cu2+, Hg2+, SDS, and p-chloromercuribenzoate. The apparent Michaelis constants of the β-xylosidase were 2.0 and 23.8 mM for p-nitrophenyl-β-xyloside and xylobiose, respectively, and those of the β-glucosidase were 1.4, 11.4, and 24.8 mM for p-nitrophenyl-β-glucoside, gentiobiose, and cellobiose, respectively. To produce xylose from crude xylooligosac-charides prepared by steam-explosion of cotton seed waste (DP ≤10, 53%, total sugars = 150 g/ liter), the crude enzyme from A. fumigatus (β-xylosidase activity = 14.7 units/ml, xylanase activity = 20 units/ml) could hydrolyze the substrate at 55°C and pH 4.5 resulting in almost complete conversion to xylose (160 g/liter). 相似文献
19.
Amaro-Reyes A García-Almendárez BE Vázquez-Mandujano DG Amaya-Llano S Castaño-Tostado E Guevara-González RG Loera O Regalado C 《Journal of industrial microbiology & biotechnology》2011,38(9):1311-1319
Xylan constitutes the second most abundant source of renewable organic carbon on earth and is located in the cell walls of
hardwood and softwood plants in the form of hemicellulose. Based on its availability, there is a growing interest in production
of xylanolytic enzymes for industrial applications. β-1,4-xylan xylosidase (EC 3.2.1.37) hydrolyses from the nonreducing end
of xylooligosaccharides arising from endo-1,4-β-xylanase activity. This work reports the partial characterization of a purified
β-xylosidase from the native strain Aspergillus niger GS1 expressed by means of a fungal system. A gene encoding β-xylosidase, xlnD, was successfully cloned from a native A. niger GS1 strain. The recombinant enzyme was expressed in A. niger AB4.1 under control of A. nidulans
gpdA promoter and trpC terminator. β-xylosidase was purified by affinity chromatography, with an apparent molecular weight of 90 kDa, and showed
a maximum activity of 4,280 U mg protein−1 at 70°C, pH 3.6. Half-life was 74 min at 70°C, activation energy was 58.9 kJ mol−1, and at 50°C optimum stability was shown at pH 4.0–5.0. β-xylosidase kept residual activity >83% in the presence of dithiothreitol
(DTT), β-mercaptoethanol, sodium dodecyl sulfate (SDS), ethylenediaminetetraacetate (EDTA), and Zn2+. Production of a hemicellulolytic free xylosidase showed some advantages in applications, such as animal feed, enzymatic
synthesis, and the fruit-juice industry where the presence of certain compounds, high temperatures, and acid media is unavoidable
in the juice-making process. 相似文献
20.
B C Saha 《Journal of industrial microbiology & biotechnology》2001,27(4):241-245
An extracellular β-xylosidase from a newly isolated Fusarium verticillioides (NRRL 26518) was purified to homogeneity from the culture supernatant by concentration by ultrafiltration using a 10,000
cut-off membrane, ammonium sulfate precipitation, DEAE Bio-Gel A agarose column chromatography and SP-Sephadex C-50 column
chromatography. The purified β-xylosidase (specific activity, 57 U/mg protein) had a molecular weight (mol. wt.) of 94,500
and an isoelectric point at pH 7.8. The optimum temperature and pH for action of the enzyme were 65°C and 4.5, respectively.
It hydrolyzes xylobiose and higher xylooligosaccharides but is inactive against xylan. The purified β-xylosidase had a K
m value of 0.85 mM (p-nitrophenol-β-D-xyloside, pH 4.5, 50°C) and was competitively inhibited by xylose with a K
i value of 6 mM. It did not require any metal ion for activity and stability. Journal of Industrial Microbiology & Biotechnology (2001) 27, 241–245.
Received 20 May 2001/ Accepted in revised form 06 July 2001 相似文献