• Asian-Australasian Journal of Animal Sciences
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• v.16 no.10
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• pp.1501-1509
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• 2003

The simultaneous addition of xylanase (5,600 EXU/kg) and phytase (500 FTU/kg) feed enzymes to wheat-based broiler diets was investigated. Starter, grower and finisher diets, with three tiers of nutrient specifications, were fed to 1,440 broiler chicks kept on deep litter from 1-42 days post-hatch, without and with xylanase plus phytase, to determine the effects of diet type and enzyme supplementation on growth performance. The nutrient specifications of type A diets were standard; energy density and protein/amino acid levels were reduced on a least-cost basis to formulate type B diets and further reduced to type C diets. Phosphorus (P) and calcium (Ca) levels were adjusted in supplemented diets. From 1-42 days post-hatch, diet type significantly influenced growth performance. Birds on type C diets had lower growth rates (2,429 vs. 2,631 g/bird; p<0.001), higher feed intakes (4,753 vs. 4,534 g/bird; p<0.005) and less efficient feed conversion (1.96 vs. 1.72; p<0.001) than birds offered type A diets. Enzyme supplementation increased growth rates by 3.2% (2,580 vs. 2,501 g/bird; p<0.005) and improved feed efficiency by 2.7% (1.80 vs. 1.85; p<0.05) over the entire feeding period. There were no interactions between diet type and enzyme supplementation. At 21 days, 5 out of 30 birds per pen were transferred to cages to ascertain treatment effect on apparent metabolisable energy (AME) and nitrogen (N) retention. Xylanase plus phytase enhanced AME (13.48 to 13.91 MJ/kg DM; p<0.001) and N retention (56.3 to 59.7%; p<0.005). Carcass and breast weights of the caged birds were determined following commercial processing. Diet type significantly influenced breast weight, carcass weight and yield. Birds offered Type A diets, in comparison to Type C diets, supported heavier breast (467 vs. 424 g; p<0.001) and carcass weights (1,868 vs. 1,699 g; p<0.001) with superior carcass yields (71.8 vs. 70.6%; p<0.005). Enzyme addition increased carcass weight by 3.9% (1,752 vs. 1,821 g; p<0.005) and breast weight by 5.8% (431 vs. 456 g; p<0.01) without influencing yields. Feed ingredient costs per kg live weight gain and per kg carcass weight indicated that enzyme addition was economically feasible, where supplementation of Type A diets generated the most effective results. Importantly, soluble and total non-starch polysaccharide and phytate contents of the wheat used were typical by local standards. This study confirms the potential of supplementing wheat-based broiler diets with xylanase plus phytase but further investigations are required to define the most appropriate inclusion rates and dietary nutrient specifications in this context.

• Asian-Australasian Journal of Animal Sciences
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• v.26 no.1
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• pp.50-58
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• 2013

A facultative bacterium producing cellulolytic and hemicellulolytic enzymes was isolated from the rumen of a native Korean goat. The bacterium was identified as a Bacillus licheniformis on the basis of biochemical and morphological characteristics and 16S rDNA sequences, and has been designated Bacillus licheniformis JK7. Endoglucanase activities were higher than those of ${\beta}$-glucosidase and xylanase at all temperatures. Xylanase had the lowest activity among the three enzymes examined. The optimum temperature for the enzymes of Bacillus licheniformis JK7 was $70^{\circ}C$ for endoglucanase (0.75 U/ml) and $50^{\circ}C$ for ${\beta}$-glucosidase and xylanase (0.63 U/ml, 0.44 U/ml, respectively). All three enzymes were stable at a temperature range of 20 to $50^{\circ}C$. At $50^{\circ}C$, endoglucanse, ${\beta}$-glucosidase, and xylanase had 90.29, 94.80, and 88.69% residual activity, respectively. The optimal pH for the three enzymes was 5.0, at which their activity was 1.46, 1.10, and 1.08 U/ml, respectively. The activity of all three enzymes was stable in the pH range of 3.0 to 6.0. Endoglucanase activity was increased 113% by $K^+$, while $K^+$, $Zn^+$, and tween 20 enhanced ${\beta}$-glucosidase activity. Xylanase showed considerable activity even in presence of selected chemical additives, with the exception of $Mn^{2+}$ and $Cu^{2+}$. The broad range of optimum temperatures (20 to $40^{\circ}C$) and the stability under acidic pH (4 to 6) suggest that the cellulolytic enzymes of Bacillus licheniformis JK7 may be good candidates for use in the biofuel industry.

• Journal of Microbiology and Biotechnology
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• v.25 no.5
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• pp.662-671
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• 2015

To enrich the genetic resource of microbial xylanases with high activity and stability under alkaline conditions, a xylanase gene (xynSL4) was cloned from Planococcus sp. SL4, an alkaline xylanase-producing strain isolated from the sediment of soda lake Dabusu. Deduced XynSL4 consists of a putative signal peptide of 29 residues and a catalytic domain (30-380 residues) of glycosyl hydrolase family 10, and shares the highest identity of 77% with a hypothetical protein from Planomicrobium glaciei CHR43. Phylogenetic analysis indicated that deduced XynSL4 is closely related with thermophilic and alkaline xylanases from Geobacillus and Bacillus species. The gene xynSL4 was expressed heterologously in Escherichia coli and the recombinant enzyme showed some superior properties. Purified recombinant XynSL4 (rXynSL4) was highly active and stable over the neutral and alkaline pH range from 6 to 11, with maximum activity at pH 7 and more than 60% activity at pH 11. It had an apparent temperature optimum of 70℃ and retained stable at this temperature in the presence of substrate. rXynSL4 was highly halotolerant, retaining more than 55% activity with 0.25-3.0 M NaCl and was stable at the concentration of NaCl up to 4M. The enzyme activity was significantly enhanced by β-mercaptoethanol and Ca²⁺ but strongly inhibited by heavy-metal ions and SDS. This thermophilic and alkaline- and salt-tolerant enzyme has great potential for basic research and industrial applications.

• Microbiology and Biotechnology Letters
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• v.48 no.2
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• pp.215-222
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• 2020

The marine microorganism PX-1, which can hydrolyze xylan, was isolated from coastal sea water of Jeju Island, Korea. Based on the 16S rRNA gene sequence and chemotaxonomy analysis, PX-1 was identified as a species of the genus Aestuariibacter and named Aestuariibacter sp PX-1. From the culture broth of PX-1, an extracellular xylanase was purified to homogeneity through ammonium sulfate precipitation and subsequent adsorption chromatography using insoluble xylan. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography estimated the molecular weight of the purified putative xylanase (XylA) as approximately 64 kDa. XylA showed xylanase activity toward beechwood xylan, with a maximum enzymatic activity at pH 6.0 and 45℃. Through thin-layer chromatographic analysis of the xylan hydrolysate produced by XylA, it was confirmed that XylA is an endo-type xylanase that decomposes xylan into xylose and xyloligosaccharides of various lengths. The K_m and V_max values of XylA for beechwood xylan were 27.78 mM and 78.13 μM/min, respectively.

• Mycobiology
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• v.33 no.2
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• pp.84-89
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• 2005

Five types of agricultural wastes were used for the production of xylanolytic enzyme by Aspergillus flavus K-03. All wastes materials supported high levels of xylanase and ${\beta}-xylosidase$ production. A high level of proteolytic activity was observed in barley and rice bran cultures, while only a weak proteolytic activity was detected in corn cob, barley and rice straw cultures. Maximum production of xylanase was achieved in basal liquid medium containing rice barn as carbon source for 5 days of culture at pH 6.5 and $25^{\circ}C$. The xylanolytic enzyme of A. flavus K-03 showed low thermostability. The times required for 50% reduction of the initial enzyme activity were 90 min at $40^{\circ}C$, 13 min at $50^{\circ}C$, and 3 min at $60^{\circ}C$. Xylanolytic activity showed the highest level at pH $5.5{\sim}10.5$ and more than 70% of the original activity was retained at pH 6.5 and 7.0. The higher stability of xylanolytic enzymes in the broad range of alkaline pH is useful for utilization of the enzymes in industrial process requiring in alkaline conditions. Moreover, the highest production of xylanolytic enzyme was obtained when 0.5% of rice bran was supplied in basal liquid medium. SDS-PAGE analysis revealed a single xylanase band of approximately 28.5 kDa from the culture filtrates.

• 한국생물공학회:학술대회논문집
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• 2001.11a
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• pp.529-532
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• 2001

Korean food waste, one of the abundantly available but environmentally problematic organic wastes in Korea, was utilized as solid-substrate by fungal strain Aspergillus niger ATcC 6275 for the production of enzymemixture containing amylase, cellulase and xylanase. The enzyme mixture can be used as high value-added animal feed. Solid-state fermentation method yielded a 84-fold enhancement in xylanase activity compared with submerged fermentation method. The effect of incubation period, incubation temperature, pH of medium, moisture content, inoculum size and enrichment of the medium with nitrogen and carbon sources were observed for optimal production of these enzymes The optimal amylase activity of 33.10 U/g, cellulase activity of 24.41 U/g, xylanase activity of 328.84 U/g were obtained at 8 days incubation with 50%(w/w) soy bean flake, with incubation temperature of $25^{\circ}C$, pH of 6.38, optimal moisture content of 55% and with inoculum size of $3.8{\times}10^6$spore/g. Enzyme activities were enhanced when ImM $CaSO_4$, 2% Malt extract and 2% galactose were added as mineral, nitrogen and carbon enrichment respectively.

• Journal of Microbiology and Biotechnology
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• v.16 no.1
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• pp.57-63
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• 2006

We have cloned a xylanase gene (xynA) from Streptomyces thermocyaneoviolaceus. The deduced amino acid sequences of the XynA, including the active site sequences of glycosyl hydrolase family 10, showed high sequence homology with several xylanases assigned in this category. The XynA was overexpressed under an IPTG inducible T7 promoter control in E. coli BLR(DE3). The overproduced enzymes were excreted into culture supernatants and periplasmic space. The purified XynA had an apparent molecular mass of near 54 kDa, which corresponds to the molecular mass calculated from its gene. The optimum pH and temperature of the purified XynA were determined to be 5.0 and $65^{\circ}C$, respectively. The XynA retained over $90\%$ its activity after the heat treatment at $65^{\circ}C$ for 30 min. The XynA was highly efficient in producing xylose (X1), xylobiose (X2), xylotriose (X3), and xylotetraose (X4) from xylan.

• Microbiology and Biotechnology Letters
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• v.24 no.1
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• pp.50-58
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• 1996

Interspecific hybrids between Aspergillus oryzae var oryzae and A. nidulans 514 were obtained by nuclear transfer technique. Several autotrophic mutants isolated from conidiospores of the two strains were mutagenized with ultraviolet and N-methyl-N-nitrosoguanidine. Optimal conditions for formation of intergeneric hybrids were investigated. Frequencies of hybrid formation by nuclear transfer were $3{\times}10^{-5}{\sim}1{\times}10^{-5}$. From observation of genetic stability, conidial size, DNA content, and nuclear stain, it was suggested that their karyptypes are aneuploid. The hybrids showed 1.1~1.4 fold higher xylanase activities than parental strains did. The xylanase of Aspergiilus sp. TAVD514-3 was purified and some of it's enzymatc characteristics were investigated. The enzyme was purified about 85 fold with an overall yield of 17% from the culture medium by ammonium sulfate fractionation, Sephadex G-75 gel permeation chromatography, and CM-sephadex A-50 ion exchange chromatography. The purified enzyme functions optimally at pH 9.0 and 80$^{\circ}C$. The enzymatic activity was increased by the presence of $Mg^{2+}$ and $Mn^2$ ions.

• Journal of Microbiology and Biotechnology
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• v.22 no.7
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• pp.930-938
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• 2012

High levels of xylanase activity (143.98 IU/ml) produced by the newly isolated Paenibacillus campinasensis G1-1 were detected when it was cultivated in a synthetic medium. A thermostable xylanase, designated XynG1-1, from P. campinasensis G1-1 was purified to homogeneity by Octyl-Sepharose hydrophobic-interaction chromatography, Sephadex G75 gel-filter chromatography, and Q-Sepharose ion-exchange chromatography, consecutively. By multistep purification, the specific activity of XynG1-1 was up to 1,865.5 IU/mg with a 9.1-fold purification. The molecular mass of purified XynG1-1 was about 41.3 kDa as estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Sequence analysis revealed that XynG1-1 containing 377 amino acids encoded by 1,134 bp genomic sequences of P. campinasensis G1-1 shared 96% homology with XylX from Paenibacillus campinasensis BL11 and 77%~78% homology with xylanases from Bacillus sp. YA-335 and Bacillus sp. 41M-1, respectively. The activity of XynG1-1 was stimulated by $Ca^{2+}$, $Ba^{2+}$, DTT, and ${\beta}$-mercaptoethanol, but was inhibited by $Ni^{2+}$, $Fe^{2+}$, $Fe^{3+}$, $Zn^{2+}$, SDS, and EDTA. The purified XynG1-1 displayed a greater affinity for birchwood xylan, with an optimal temperature of $60^{\circ}C$ and an optimal pH of 7.5. The fact that XynG1-1 is cellulose-free, thermostable (stability at high temperature of $70^{\circ}C{\sim}80^{\circ}C$), and active over a wide pH range (pH 5.0~9.0) suggests that the enzyme is potentially valuable for various industrial applications, especially for pulp bleaching pretreatment.

• Journal of Microbiology and Biotechnology
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• v.24 no.11
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• pp.1559-1565
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• 2014

Cellulase and xylanase are main hydrolysis enzymes for the degradation of cellulosic and hemicellulosic biomass, respectively. In this study, our aim was to develop and test the efficacy of a rapid, high-throughput method to screen hydrolytic-enzyme-producing microbes. To accomplish this, we modified the 3,5-dinitrosalicylic acid (DNS) method for microwell plate-based screening. Targeted microbial samples were initially cultured on agar plates with both cellulose and xylan as substrates. Then, isolated colonies were subcultured in broth media containing yeast extract and either cellulose or xylan. The supernatants of the culture broth were tested with our modified DNS screening method in a 96-microwell plate, with a $200{\mu}l$ total reaction volume. In addition, the stability and reliability of glucose and xylose standards, which were used to determine the enzymatic activity, were studied at $100^{\circ}C$ for different time intervals in a dry oven. It was concluded that the minimum incubation time required for stable color development of the standard solution is 20 min. With this technique, we successfully screened 21 and 31 cellulase- and xylanase-producing strains, respectively, in a single experimental trial. Among the identified strains, 19 showed both cellulose and xylan hydrolyzing activities. These microbes can be applied to bioethanol production from cellulosic and hemicellulosic biomass.