• Journal of Microbiology and Biotechnology
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• v.31 no.8
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• pp.1123-1133
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• 2021

Biodegradation is the key process involved in natural lignocellulose biotransformation and utilization. Microbial consortia represent promising candidates for applications in lignocellulose conversion strategies for biofuel production; however, cooperation among the enzymes and the labor division of microbes in the microbial consortia remains unclear. In this study, metagenomic analysis was performed to reveal the community structure and extremozyme systems of a lignocellulolytic microbial consortium, TMC7. The taxonomic affiliation of TMC7 metagenome included members of the genera Ruminiclostridium (42.85%), Thermoanaerobacterium (18.41%), Geobacillus (10.44%), unclassified_f__Bacillaceae (7.48%), Aeribacillus (2.65%), Symbiobacterium (2.47%), Desulfotomaculum (2.33%), Caldibacillus (1.56%), Clostridium (1.26%), and others (10.55%). The carbohydrate-active enzyme annotation revealed that TMC7 encoded a broad array of enzymes responsible for cellulose and hemicellulose degradation. Ten glycoside hydrolases (GHs) endoglucanase, 4 GHs exoglucanase, and 6 GHs β-glucosidase were identified for cellulose degradation; 6 GHs endo-β-1,4-xylanase, 9 GHs β-xylosidase, and 3 GHs β-mannanase were identified for degradation of the hemicellulose main chain; 6 GHs arabinofuranosidase, 2 GHs α-mannosidase, 11 GHs galactosidase, 3 GHs α-rhamnosidase, and 4 GHs α-fucosidase were identified as xylan debranching enzymes. Furthermore, by introducing a factor named as the contribution coefficient, we found that Ruminiclostridium and Thermoanaerobacterium may be the dominant contributors, whereas Symbiobacterium and Desulfotomaculum may serve as "sugar cheaters" in lignocellulose degradation by TMC7. Our findings provide mechanistic profiles of an array of enzymes that degrade complex lignocellulosic biomass in the microbial consortium TMC7 and provide a promising approach for studying the potential contribution of microbes in microbial consortia.

• Korean Journal of Soil Science and Fertilizer
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• v.37 no.3
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• pp.165-170
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• 2004

This study was conducted to investigate the effect of different surface soil management practices on soil microflora in volcanic ash soils of citrus orchard. Soil samples were collected from citrus orchards of clean cultivation, grass sod, and grass mulch system in May and September 1997. Soil chemical properties, populations of various microorganisms, enzyme activities, microbial biomass C were analyzed. Average soil pH were 4.7, and average nitrogen and organic matter contents were 6 and $140.2g\;kg^{-1}$, respectively. Aerobic bacteria were distributed at $26,2-47.3{\times}10^6cfu\;g^{-1}$ level. Among the aerobic bacteria Pseudomonas spp., Rhizobium spp., and thermophilic Bacillus spp. were dominant in most of the investigated orchard soils. Density of actinomycetes were low at $1.8-84.6{\times}10^5cfu\;g^{-1}$ level. Fungi were distributed at $26.4-182.1{\times}10^5cfu\;g^{-1}$ level and the density was higher in grass mulch and sward sites. In september, phosphomonoesterase activity was high at $239.6{\mu}g\;PNP\;g\;soil^{-1}\;h^{-1}$ in clean cultivated citrus orchards. Soil cellulase activity were higher at $602.6{\mu}g\;GE\;g\;soil^{-1}$\;24\;h^{-1}$ in grass sward cultivation than any other soil management practices. Soil microbial biomass C was higher in grass mulch cultivated orchards.

• Journal of Animal Science and Technology
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• v.50 no.5
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• pp.713-720
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• 2008

This study was conducted to isolate and identify xylanase- and cellulase-producing thermophilic bacteria from stacked spent mushroom substrates and to determine the optimal medium conditions for their growth. Bacteria with the highest xylanase and CMCase activities were strain 3 and 201-7. Both of them were identified as Bacillus spp. and named Bacillus subtilis KU3 and Bacillus subtilis KU201-7. The optimal medium condition of Bacillus subtilis KU3 was obtained when 3%(w/v) of yeast extract and 1%(w/v) of maltose were used as nitrogen and carbon sources, respectively. That of Bacillus subtilis KU201-7 was obtained when 0.5%(w/v) of yeast extract and 0.5%(w/v) of CMC were used as nitrogen and carbon sources, respectively.

• KSBB Journal
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• v.10 no.3
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• pp.304-316
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• 1995

For fuel alcohol production, enzymatic liquefaction and saccharification of tapioca starch by ${\alpha}$-amylase and glucoamylase were studied. The thermophilic ${\alpha}$-amylase Termamyl produced from Bacillus licheniformis gave a better liquefaction than the relalively low temperature enzyme BAN from B. subtilis. Oplimal temperature and pH with Termamyl were $90∼95^{\circ}C$ and 5.8, respectively. Minimal amount of Termamyl 240uc for a satisfactory liquefaction for a two-hour reaction was about 0.0125% (v/w) with respect to the mass of tapioca used. For saccharification experiments two enzymes, Novo AMG and Do-I1 enzymes were compared. The enzymatic activity of each enzyme was a little different depending on the substrate used and the latter was found to have a significant amount of ${\alpha}$-amylase activity. With Novo AMG optimal temperature was about $58^{\circ}C$ The pH optimum was 4.3 with maltose, however, with tapioca, no difference was observed between pH 4.3 and 5.7 which is a natural, unadjusted pH of liquefied tapioca. For 85% of completion of saccharification, it was necessary to use 0.0625% (v/w) of Novo AMG 400L for tapioca and to run the reaction for more than 10 hr, Packed volume of solid particles in tapioca slurry remained at around 30% during liquefaction and saccharification. This indicates that the removal of the solid particle before fermentation is not economically feasible at all, even though the solid particles make it very difficult to operate the bioreactor in a continuous mode with cell-recycle.

• Journal of Microbiology and Biotechnology
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• v.22 no.4
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• pp.501-509
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• 2012

A 990 bp full-length gene (xynAHJ2) encoding a 329-residue polypeptide (XynAHJ2) with a calculated mass of 38.4 kDa was cloned from Bacillus sp. HJ2 harbored in a saline soil. XynAHJ2 showed no signal peptide, distinct amino acid stretches of glycoside hydrolase (GH) family 10 intracellular endoxylanases, and the highest amino acid sequence identity of 65.3% with the identified GH 10 intracellular mesophilic endoxylanase iM-KRICT PX1-Ps from Paenibacillus sp. HPL-001 (ACJ06666). The recombinant enzyme (rXynAHJ2) was expressed in Escherichia coli and displayed the typical characteristics of low-temperature-active enzyme (exhibiting optimum activity at $35^{\circ}C$, 62% at $20^{\circ}C$, and 38% at $10^{\circ}C$; thermolability at ${\geq}45^{\circ}C$). Compared with the reported GH 10 low-temperature-active endoxylanases, which are all extracellular, rXynAHJ2 showed low amino acid sequence identities (<45%), low homology (different phylogenetic cluster), and difference of structure (decreased amount of total accessible surface area and exposed nonpolar accessible surface area). Compared with the reported GH 10 intracellular endoxylanases, which are all mesophilic and thermophilic, rXynAHJ2 has decreased numbers of arginine residues and salt bridges, and showed resistance to $Ni^{2+}$, $Ca^{2+}$, or EDTA at 10 mM final concentration. The above mechanism of structural adaptation for low-temperature activity of intracellular endoxylanase rXynAHJ2 is different from that of GH 10 extracellular low-temperature-active endoxylanases. This is the first report of the molecular and biochemical characterizations of a novel intracellular low-temperature-active xylanase.

• Microbiology and Biotechnology Letters
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• v.2 no.3
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• pp.133-140
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• 1974

1) Two xylanase (designed as A and B) of Myriococcus albomyces were purified from an extract of wheat koji culture. Puriscation steps included first ammonium sulfate fractionation followed successively by SE-Sephadex column chromatgraphy. DEAE-Sephadex column chromatography and gel filtration on Sephadex G-100 repectively. 2) The optimum pH and pH stability for crude xylanse were found to be pH 5.0 and pH 4.0-7.0 respectively. 3) The optimium temperature was found to he 5$0^{\circ}C$ and for the thermal statbility of xylanase, the enryme incubated at $65^{\circ}C$ for 60min did not affect their stability. 4) The purised xylanase A and B were considered as liquefying xylanase and saccharogenic xylauase repectively. 5) The Bylanase A was most active at pH 4.0 and range of pH 3.0-8.0 at 3$0^{\circ}C$ for six hrs. The B was most active at pH 5.0 showing stability range of pH 4.0 to 8.5 at 3$0^{\circ}C$ for 6 hrs. incubation respectively. The Optimum temperature of xylanase A and B were found to be 7$0^{\circ}C$ and $65^{\circ}C$ for 60min repectively.

• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.730-738
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• 2016

est19 is a gene from Bacillus sp. K91 that encodes a new esterase. A comparison of the amino acid sequence showed that Est19 has typical Ser-Gly-Asn-His (SGNH) family motifs and could be grouped into the SGNH hydrolase family. The Est19 protein was functionally cloned, and expressed and purified from Escherichia coli BL21(DE3). The enzyme activity was optimal at 60℃ and pH 9.0, and displayed esterase activity towards esters with short-chain acyl esters (C₂-C₆). A structural model of Est19 was constructed using phospholipase A1 from Streptomyces albidoflavus NA297 as a template. The structure showed an α/β-hydrolase fold and indicated the presence of the typical catalytic triad Ser49-Asp227-His230, which were further investigated by site-directed mutagenesis. To the best of our knowledge, Est19 is a new member of the SGNH hydrolase family identified from thermophiles, which may be applicable in the industrial production of semisynthetic β-lactam antibiotics after modification.