• 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.

• 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.16 no.7
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• pp.1132-1138
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• 2006

Three types of xylanases (EC 3.2.1.8) were detected in the strain Aspergillus niger A-25, one of which, designated as XynIII, also displayed ${\beta}-(l,3-1,4)-glucanase$ (EC 3.2.1.73) activity, as determined by a zymogram analysis. XynIII was purified by ultrafiltration and ion-exchange chromatography methods. Its apparent molecular weight was about 27.9 kDa, as estimated by SDS-PAGE. The purified XynIII could hydrolyze birchwood xylan, oat spelt xylan, lichenin, and barley ${\beta}-glucan$, but not CMC, avicel cellulose, or soluble starch under the assay conditions in this study. The xylanase and ${\beta}-(l,3-1,4)-glucanase$ activities of XynIII both had a similar optimal pH and pH stability, as well as a similar optimal temperature and temperature stability. Moreover, the effects of metal ions on the two enzymatic activities were also similar. The overall hydrolytic rates of XynIII in different mixtures of xylan and lichenin coincided with those calculated using the Michaelis-Menten model when assuming the two substrates were competing for the same active site in the enzyme. Accordingly, the results indicated that XynIII is a novel bifunctional enzyme and its xylanase and ${\beta}-(l,3-1,4)-glucanase$ activities are catalyzed by the same active center.

• The Korean Journal of Mycology
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• v.42 no.1
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• pp.86-90
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• 2014

Button mushroom (Agaricus bisporus) strains from diverse origins were compared in this study to obtain basic information on their growth and biochemical properties that are helpful for breeding. Among 31 dikaryotic strains tested, most strains showed better mycelial growth on oatmeal agar than on MEA and PDA. Mycelia of the mushroom strains revealed ${\beta}$-glucosidase activity the most clearly among the seven extracellular enzymes tested. All the strains showed protease activity, but ${\beta}$-glucosidase activity was found in 27 strains and xylanase activity was found in 30 strains. The activity of avicelase, CM-cellulase, amylase, and pectinase was detected in less than 20 strains. These results implied that enzymatic characteristics could be used as a criterion of strain selection for breeding study.

• Korean Journal of Microbiology
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• v.48 no.2
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• pp.73-78
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• 2012

We previously reported that the truncated Cel44C-$Man26A_{P558}$ ${\beta}$-glycosyl hydrolase protein exhibits multifunctional activities, including cellulase, xylanase, and lichenase. DNA shuffling of the truncated Cel44C-$Man26A_{P558}$ enzyme was performed to enhance the enzymatic activity of the multifunctional ${\beta}$-glycosyl hydrolase. Two mutant enzymes, M2Cel44C-$Man26A_{P558}$ that carries one mutation (P438A) and M21Cel44C-$Man26A_{P558}$ that carries two mutations (A273T and P438A) were obtained. The enzymatic activity of the M21Cel44C-$Man26A_{P558}$ double mutant was lower than enzymatic activity of the single mutant (M2Cel44C-$Man26A_{P558}$). However, both mutants displayed the enhancements in their enzyme activities that were ${\approx}1.3$- to 2.2-fold higher than the original enzymatic activity in Cel44C-$Man26A_{P558}$. In particular, the mutant M2Cel44C-$Man26A_{P558}$ exhibited an approximate 1.5- to 2.2-fold increase in the cellulase, xylanase, and lichenase activities in comparison with the control (Cel44C-$Man26A_{P558}$). The optimum cellulase, linchenase, and xylanase activities of ${\beta}$-glycosyl hydrolase were observed at pH 7.0, pH 7.0 and pH 6.0, respectively. These results, therefore, suggest that the amino acid residue Ala438 plays important roles in the enhancement of the activity of multifunctional ${\beta}$-glycosyl hydrolase.

• Journal of Animal Science and Technology
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• v.46 no.5
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• pp.763-772
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• 2004

The patterns of fungal growth and fiber digestion under the microscope, and tile productions of fibrolytic enzyme were studied in an in vitro culture with Neocallimastix frontalis SA when either filter paper or rice straw was provided as sole energy source. Under the microscopic observation, active zoospores attachment, sporangium development and complex rhizoidal system were founded on the surface and at the edge of filter paper. After 7 days of incubation, a reduced fiber mass, a decreased fiber cohesion and a weakened fiber structure by fungal digestion were clearly observed. Similar fungal development was observed with rice straw, but fungal growth and digestion took place mostly on the damaged and exposed portion of rice straw. Although there were some differences in absolute concentration and pattern, the concentration of both cellulase and xylanase increased with incubation time with the higher activity being obtained with filter paper. Their differences were large especially after 48 and 96hr of incubation(P< 0.05). The filter paper was more good inducer of cellulolytic and xylanolytic enzymes compared with complex substrate, rice straw. These findings suggest that the filter paper is the better energy source for N frontalis than the complex substrate, and structural disintegration by physical process is able to help rumen fungal growth on the lignified roughage although anaerobic rumen fungi have mechanical and enzymatic functions for fiber digestion.

• Current Research on Agriculture and Life Sciences
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• v.16
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• pp.45-54
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• 1998

A thermotolerant bacterium, Streptomyces thermocyaneoviolaceus which produced xylan-degrading enzymes, utilized excellently xylan of wheat bran by producing the enzymes in comparison with that of birchwood or oat spelts. Optimal enzyme production was achieved in WB medium containing 0.8% wheat bran, 0.06% yeast extract, 0.06% bactopeptone, 0.05% $MgSO_4{\cdot}7H_2O$, 0.05% $FeSO_4{\cdot}7H_2O$, 0.05% $KH_2PO_4$ and, 0.2% $K_2HPO_4$(pH 7.0) at $50^{\circ}C$ for 24 hrs. The optimal pH and temperature for the hydrolysis of xylan were pH 5.5 and $65^{\circ}C$, respectively. The enzyme activity was retained more than 80% at the range from pH 4.5 to pH 9.5 at $4^{\circ}C$ for 12 hrs and 94% on the heat-treatment at $65^{\circ}C$ for 1 hr. Xylobiose, xylotriose, xylose, and other xylooligosaccharides were produced as end products from hydrolysis of birchwood xylan by the xylanase of S. thermocyaneoviolceus.

• Journal of Microbiology and Biotechnology
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• v.9 no.6
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• pp.873-876
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• 1999

Aspergillus sp. CX-l strain grown on microcrystalline cellulose resulted in the accumulation of high levels of cellulase and xylanase activities that were higher by two to four folds than those from the conventional commercial producer, Trichoderma reesei QM9414. Aspergillus sp. CX-1 demonstrated greater thermo stability and better catalytic characteristics of total cellulase activity (FPase) as compared to T. reesei and Aspergillus niger F-2039.

• KSBB Journal
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• v.32 no.2
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• pp.96-102
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• 2017

The lignocellulose that is a major component of spent coffee ground was degraded and saccharified. To implement the spent coffee, after several pre-treatments, inoculation of Phanerochaete chrysosporium and solid-state fermentation were conducted. The optimal temperature of the enzymes (lignin peroxidase, manganese peroxidase, xylanase, laccase, and cellulase) for degradation of lignocellulose by P. chrysosporium was found. We also measured the maximum activity of enzymes (lignin peroxidase 0.15 IU/mL, manganese peroxidase 0.90 IU/mL, laccase 0.11 IU/mL, cellulase 5.87 IU/mL, carboxymethyl cellulase 9.52 IU/mL, xylanase 1.16 IU/mL) used for the process. As a result, 4.73 mg/mL of reduced sugar was obtained and 61.02% of lignin was degraded by solid state fermentation of P. chrysosporium on spent coffee ground.

• 한국생물공학회:학술대회논문집
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• 2001.11a
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• pp.629-632
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• 2001

The strain FJ1 isolated from a rotten wood showed high activity to hydrolysis of cellulosic materials. The strain produced largely enzymes related in hydrolysis of cellulose and hemicellulose, such as CMCase, xylanase, ${\beta}-glucosidase$ , and avicelase. The culture conditions(pH, temperature, inoculation concentration) and substrate specificity to various cellulosic materials were examined to elevate productivity of the enzymes. The enzyme activities of CMCase and xylanase were 13.5U/ml and 24.3U/ml in agitation culture using Mandel's medium, respectively. The high activity of the enzymes was earned when mixed cellulosic materials of rice straw, sawdust, and pulp as substrates, indicating that the strain FJ1 could use crystalline substrates.