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1.
Production and Properties of Xylan-Degrading Enzymes from Cellulomonas uda 总被引:2,自引:2,他引:0
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Xylan degradation and production of β-xylanase and β-xylosidase activities were studied in cultures of Cellulomonas uda grown on purified xylan from birchwood. β-Xylanase activity was found to be associated with the cells, although in various degrees. The formation of β-xylanase activity was induced by xylotriose and repressed by xylose. β-Xylosidase activity was cell bound. Both constitutive and inducible β-xylosidase activities were suggested. β-Xylanase and β-xylosidase activities were inhibited competitively by xylose. β-Xylanase activity had a pronounced optimum pH of 5.8, whereas the optimum pH of β-xylosidase activity ranged from 5.4 to 6.1. The major products of xylan degradation by a crude preparation of β-xylanase activity, in decreasing order of amount, were xylobiose, xylotriose, xylose, and small amounts of xylotetraose. This pattern suggests that β-xylanase activity secreted by C. uda is of the endosplitting type. Supernatants of cultures grown on cellulose showed not only β-glucanase but also β-xylanase activity. The latter could be attributed to an endo-1,4-β-glucanase activity which had a low β-xylanase activity. 相似文献
2.
Abstract Fusarium oxysporum f. sp. melonis produces extracellular endo-1,4-β-xylanase and β-xylosidase when grown in shaken culture at 26°C in a mineral salts medium containing oat spelt xylan and glucose as carbon sources. Endo-1,4-β-xylanase was purified 251 times from 5-day-old culture filtrates, by Sephacryl S-200, ion exchange and gel filtration HPLC. The purified sample yielded a single band in SDS polyacrylamide gels with a molecular mass of 80 kDa on electrophoretic mobility and 83 kDa by gel filtration behavior. High activity of the endo-1,4-β-xylanase against xylan was observed between 5 and 8 pH, and between 40 and 60°C, the optimum pH and temperature being 5.0 and 50°C, respectively. Kinetic properties of the enzyme are similar to those of other fungal xylanases, showing high affinity towards oat spelt xylan with a K m of 1 mM expressed as xylose equivalent. 相似文献
3.
Here, we present the results of a computational analysis of a group of hypothetical GH10 endo-β-xylanases from the Planctomycetes, a bacterial phylum with poorly characterized functional capabilities. These proteins are encoded in all analyzed genomes of heterotrophic Planctomycetes and form a phylogenetically distinct and tight cluster. In addition, we determined nucleotide sequences for endo-β-xylanase genes from five strains of Isosphaera-Singulisphaera group of the Planctomycetes. The trees constructed for the 16S rRNA genes and the inferred amino acid sequences of endo-β-xylanases were highly congruent, thus suggesting the vertical transfer of endo-β-xylanase genes and their functional importance in Planctomycetes. 相似文献
4.
《Bioscience, biotechnology, and biochemistry》2013,77(8):1722-1724
cDNA encoding an extracellular carbohydrate esterase (CcEst1) was cloned from the basidiomycete Coprinopsis cinerea. The recombinant CcEst1 expressed in Pichia pastoris acted on p-nitrophenyl acetate, α-naphthyl acetate, and methyl hydroxycinnamic acids, except for methyl sinapic acid. The enzyme released ferulic and acetic acids from wheat arabinoxylan and acetylated xylan respectively. Activity increased on the addition of endo-β-1,4-xylanase. 相似文献
5.
Rikiya Kuwahara Tateo Suzuki Hiroshi Meguro 《Bioscience, biotechnology, and biochemistry》2013,77(5):1195-1196
A purified extracellular endo β-1,3-xylanase (EC 3.2.1.32) from an isolated strain, Aspergillus terreus A-07, was found to hydrolyze 1,3-xylosyl linkages only. When rhodymenan (β-1,4 and β-1.3-linked xylan) was hydrolyzed by β-1,3-xylanase (EF-6), four β-1,4-linked xylooligosaccharide fractions were produced. The main product was β-1,4-xylotriose, with trace amounts of other β-1,4-linked xylooligosaccharides. Successive degradation by β-l,4-xylosidase of the β,4-xylooligosaccharides that were produced from hydrolysis of β-1,3-xylanase on rhodymenan yielded only xylose as the final product.We compared the action pattern of this enzyme with that of an extracellular endo β-l,4-xylanase (EC 3.2.1.8) of Streptomyces. From a mixture of products of β-1,4-xylanase hydrolysis on rhodymenan, an isomeric xylotriose was isolated by charcoal chromatography after treating with β-1.4-xylosidase. The structure of this isomeric xylotriose was elucidated by methylation analysis and its susceptibility to β-1,4-xylanase, β-1,3-xylanase, and β-1,4-xylosidase. The obtained isomeric xylotriose was identified as 3-O-β-xylopyranosyl-4-O-β-D-xylopyranosyl-D-xylose (X1→3X1→4X). It has a melting point of 224~225°C and (c = 1, H2O)= —46°. 相似文献
6.
dos Santos Jaqueline Pozzada Dittgen Caroline Lambrecht El Halal Shanise Lisie Mello Vanier Nathan Levien 《Food biophysics》2020,15(2):155-161
Food Biophysics - A versatile and effective method of producing polyvinyl alcohol (PVA)-xylanase (XY) fibers cross-linked by glutaraldehyde vapor (GA) is reported in the present study. Crosslinking... 相似文献
7.
The Cryptococcus albidus XLN-gene (encoding endo-1,4-β-xylanase) was expressed in the yeast Pichia stipitis under the control of the PsADH2-promoter, which is activated under O2 limitation. The resulting transformant produced endo-1,4-β-xylanase after a shift to anoxic conditions. Endo-1,4-β-xylanase production was enhanced by limited aeration after the shift. 相似文献
8.
Cloning, Sequencing, and Expression in Escherichia coli of the New Gene Encoding β-1,3-Xylanase from a Marine Bacterium, Vibrio sp. Strain XY-214
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The Vibrio sp. strain XY-214 β-1,3-xylanase gene cloned in Escherichia coli DH5α consisted of an open reading frame of 1,383 nucleotides encoding a protein of 460 amino acids with a molecular mass of 51,323 Da and had a signal peptide of 22 amino acids. The transformant enzyme hydrolyzed β-1,3-xylan to produce several xylooligosaccharides. 相似文献
9.
Utilization of waste cellophane for production of extracellular cellulose-degrading enzymes byTrichoderma reesei was studied on laboratory-scale experiments using liquid media containing varnished or non-varnished cellophane disintegrated physically to different degrees. Enzyme activities were directly proportional to the degree of cellophane disintegration,i.e. its specific surface area. Hydrolytic activities of cellulase on 1 % milled cellophane reached approximately 50 % yield of the activities acquired on 1 % microcrystalline cellulose. Addition of Xylocel, concentrated prehydrolyzate of beech wood, added to the media in amounts corresponding to 0.2 to 1.0 % concentration of reducing groups, did not improve the yield of cellulase on milled cellophane. However, β-D-xylanase activity was increased several times due to the presence of lower xylooligosaccharides serving as specific β-D-xylanase inducers. No differences in occurrence of multiple forms of endo-l,4-β-D-glueanases and endo-l,4-β-D-xylanases and their isoelectric points were observed. 相似文献
10.
《Bioscience, biotechnology, and biochemistry》2013,77(6):1041-1044
To discover the specificity of Streptomyces β-xylanase toward xylan having glucose stubs, glucoxylan was prepared by the hydrogenation of cotton-seed cake glucuronoxylan. The glucoxylan was hydrolyzed by the β-xylanase of Streptomyces olivaceoviridis E-86, and three kinds of glucoxylo-oligosaccharides were isolated from the hydrolysate by chromatographies on a charcoal column, preparative paper partition, and a Toyopearl HW-40F column. The isolated oligosaccharides had the structures of 23-α-glucopyranosylxylo-triose, 23-α-(4–O-methyl-glucopyranosyl)xylotriose and 24-α-glucopyranosylxylotetraose. From the structure of the above oligosaccharides and the results of our previous studies, we suggest that the specificity of Streptomyces β-xylanase toward glucose stubs is the same as that toward glucuronic acid stubs, but differs considerably from that toward arabinose stubs. 相似文献
11.
Recently we reported the molecular cloning and characterization of a novel beta-1,3-xylanase from the marine bacterium Vibrio sp. AX-4 [Kiyohara et al. (2005) Biochem. J. 388, 949-957]. We report here the structural analysis of oligosaccharides generated from beta-1,3-xylan of a siphonous green alga, Caulerpa racemosa var. laete-virens, by the action of beta-1,3-xylanase. The enzyme degraded the polysaccharide producing oligosaccharides with different R(f)s on TLC (EX2-EX5). Sugar component, linkage, and MALDI-TOF-MS analyses revealed that EX2 and EX3 were Xyl-1,3-Xyl and Xyl-1,3-Xyl-1,3-Xyl, respectively. On the other hand, EX4 was a mixture of Glc-1,3-Xyl-1,3-Xyl, Xyl-1,4-Xyl-1,3-Xyl and Xyl-1,3-Xyl-1,4-Xyl, while EX5 was a mixture of tetra-saccharides containing 3-substitued Glc in addition to the same components of EX4. Branching was not likely present in EXOs prepared from the polysaccharide by the enzyme. These results strongly suggest that the C. racemosa beta-1,3-xylan is a linear heteropolysaccharide containing 1,3-Glc and 1,4-Xyl both of which are thought to be located within a beta-1,3-Xyl chain and linked via covalent bonds. This report indicates the usefulness of the enzyme for the structural analysis of beta-1,3-xylan. 相似文献
12.
Sequence analysis of β-1,3-xylanase (TxyA) from a marine bacterium, Alcaligenes sp. strain XY-234 implied that an xylan-binding module belonging to carbohydrate-binding module family 31 (TxyA-CBM) is separated
from a catalytic module belonging to glycosyl hydrolase family 26 (TxyA-CM) by a putative glycine-rich linker [Okazaki, F.,
et al. (2002) J. Bacteriol. 184: 2399–2403]. In order to reveal the role of these structural features of TxyA, two modules, TxyA-CBM
and TxyA-CM, were constructed, and those modules and full-length TxyA were characterized by thermodynamic studies. TxyA-CBM
and TxyA-CM showed full reversible folding from denaturant-induced unfolded forms, exhibited higher thermodynamic stabilities.
The conformational stability of both truncated modules is industrially desirable, as well as aiding the understanding of the
enzymatic characterization of the two modules of β-1,3-xylanase separated by a long linker. 相似文献
13.
《Bioscience, biotechnology, and biochemistry》2013,77(11):2068-2074
Three kinds of xylo-oligosaccharides having structures of 32-β-xylosylxylobiose, 32-β-xylobiosylxylobiose, and 22-β-xylobiosylxylobiose were isolated from an enzymatic hydrolysate of hardwood xylan with Streptomyces β-xylanase. The structures suggest that the hardwood xylan has both (1 → 2)- and (1 → 3)-β-d-xylopyranosyl linkages in the structure, and the specificity of Streptomyces β-xylanase toward the stubs is similar to that toward glucuronic acid stubs, but is somewhat different from that toward arabinose and xylosylarabinose stubs. 相似文献
14.
宇佐美曲霉木聚糖酶基因xynⅡ在不同毕赤酵母中的分泌表达 总被引:2,自引:1,他引:2
将宇佐美曲霉E001的内切-1,4-木聚糖酶基因克隆到毕赤酵母表达载体pPIC9K中,得到重组质粒pPXY-NII,将其经SalⅠ线性化后分别转化2株毕赤酵母GS115和KM71,xynⅡ基因通过同源重组被整合到毕赤酵母染色体上,并处于酵母α因子的下游,经筛选获得阳性重组菌PXGL98(Mut+)和PXKL29(Muts)。该木聚糖酶基因在2株毕赤酵母中均实现了分泌表达。同时对工程菌的发酵条件进行了优化,在甲醇诱导下,PXGL98与PXKL29培养物上清液中的酶活力分别可达1156.92 U/mL和1646.03 U/mL。 相似文献
15.
Two endo-β-1,4-xylanase-encoding genes were amplified from Aspergillus niger ATCC 90196 mRNA, inserted between the yeast ADH2
promoter and terminator sequences (genes designated XYN4 and XYN5) and expressed in Saccharomyces cerevisiae. The nucleotide
sequences of the XYN4 and XYN5 genes revealed that both genes encode 211-amino acid proteins that are 92% identical to each
other. Both the Xyn4 and Xyn5 enzymes have pH and temperature optima of pH 4 and 60°C, respectively. Autoselective S. cerevisiae
strains were developed that allowed β-xylanase production and secretion in complex medium.
This revised version was published online in November 2006 with corrections to the Cover Date. 相似文献
16.
The 1.7A resolution crystal structure of recombinant family G/11 beta-1,4-xylanase (rXynA) from Bacillus subtilis 1A1 shows a jellyroll fold in which two curved beta-sheets form the active-site and substrate-binding cleft. The onset of thermal denaturation of rXynA occurs at 328 K, in excellent agreement with the optimum catalytic temperature. Molecular dynamics simulations at temperatures of 298-328 K demonstrate that below the optimum temperature the thumb loop and palm domain adopt a closed conformation. However, at 328 K these two domains separate facilitating substrate access to the active-site pocket, thereby accounting for the optimum catalytic temperature of the rXynA. 相似文献
17.
Coprophilous fungi inhabit herbivore feces, secreting enzymes to degrade the most recalcitrant parts of plant biomass that have resisted the digestive process. Consequently, the secretomes of coprophilous fungi have high potential to contain novel and efficient plant cell wall degrading enzymes of biotechnological interest. We have used one-dimensional and two-dimensional gel electrophoresis, matrix-assisted laser desorption ionization-time-of-flight tandem mass spectrometry (MALDI-TOF/TOF MS/MS), and quadrupole time-of-flight liquid chromatography-tandem mass spectrometry (Q-TOF LC-MS/MS) to identify proteins from the secretome of the coprophilous fungus Doratomyces stemonitis C8 (EU551185) isolated from koala feces. As the genome of D. stemonitis has not been sequenced, cross-species identification, de novo sequencing, and zymography formed an integral part of the analysis. A broad range of enzymes involved in the degradation of cellulose, hemicellulose, pectin, lignin, and protein were revealed, dominated by cellobiohydrolase of the glycosyl hydrolase family 7 and endo-1,4-β-xylanase of the glycosyl hydrolase family 10. A high degree of specialization for pectin degradation in the D. stemonitis C8 secretome distinguishes it from the secretomes of some other saprophytic fungi, such as the industrially exploited T. reesei. In the first proteomic analysis of the secretome of a coprophilous fungus reported to date, the identified enzymes provide valuable insight into how coprophilous fungi subsist on herbivore feces, and these findings hold potential for increasing the efficiency of plant biomass degradation in industrial processes such as biofuel production in the future. 相似文献
18.
The xynB gene, which encodes endo-beta-1,4-xylanase XynB, in Aspergillus niger BRFM281 was amplified by RT-PCR using mRNA isolated from a culture containing sugar beet pulp as an inducer. The cDNA was cloned into an expression cassette under the control of the strong and constitutive glyceraldhehyde-3-phosphate dehydrogenase gene promoter. The expression system was designed to produce the recombinant enzyme XynB with a six-histidine peptide fused to the carboxy end of the protein. Homologous overproduction of XynB was successfully achieved in shake flask cultures, and the secretion yield was estimated to be 900 mg x L(-1). The recombinant XynB was purified 1.5-fold by immobilized metal affinity chromatography to homogeneity using a one-step purification protocol with 71% recovery. The purified recombinant enzyme was fully characterized and has a molecular mass of 23 kDa and an optimal activity at pH 5.5 and 50 degrees C with stability in the pH range 4.0-7.0 and temperature up to 50 degrees C. Using soluble oat spelts xylan, the determined Km and Vmax values were 7.1 mg x mL(-1) and 3881 U x mg(-1), respectively. 相似文献
19.
Phytopathogenic fungi can degrade xylan, an abundant hemicellulose in plant cell walls, by the coordinate action of a group of extracellular enzymes. Among these, endo-beta-1,4-xylanases carry out the initial breakdown by cleaving internal bonds in the polymer backbone. We have isolated and characterized a gene, xyn11A, coding for an endo-beta-1,4-xylanase belonging to family 11 of glycosyl hydrolases. xyn11A was shown to be induced by xylan and repressed by glucose and to be expressed in planta. The disruption of xyn11A caused only a moderate decrease, about 30%, in the level of extracellular endo-beta-1-4-xylanase activity and in the growth rate, with beechwood xylan as the only carbon source. However, deletion of the gene had a more pronounced effect on virulence, delaying the appearance of secondary lesions and reducing the average lesion size by more than 70%. Reintroducing the wild-type gene into the mutant strains reversed this phenotype back to wild type. 相似文献
20.
Two chimeric genes, XynA-Bs-Glu-1 and XynA-Bs-Glu-2, encoding Aspergillus sulphureus β-xylanase (XynA, 26 kDa) and Bacillus subtilis β-1,3-1,4-glucanase (Bs-Glu, 30 kDa), were constructed via in-fusion by different linkers and expressed successfully in Pichia pastoris. The fusion protein (50 kDa) exhibited both β-xylanase and β-1,3-1,4-glucanase activities. Compared with parental enzymes, the moiety activities were decreased in fermentation supernatants. Parental XynA and Bs-Glu were superior to corresponding moieties in each fusion enzymes because of lower Kn higher kcat. Despite some variations, common optima were generally 50°C and pH 3.4 for the XynA moiety and parent, and 40°C and pH 6.4 for the Bs-Glu counterparts. Thus, the fusion enzyme XynA-Bs-Glu-1 and XynA-Bs-Glu-2 were bifunctional. 相似文献