• Microbiology and Biotechnology Letters
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• v.32 no.3
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• pp.212-217
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• 2004

The gene coding for mannanase from Bacillus subtilis WL-7, a number of glycosyl hydrolase family 26, was hyperexpressed in Escherichia coli. Two recombinant plasmids, pE7MAN and pENS7, were constructed by introducing the complete mannanase gene and the mature mannanase gene lacking N-terminal signal peptide region into a expression vector pET24a(＋), respectively. The level of mannanase produced by E. coli BL21 (DE3) carrying pENS7, which included the mature mannanase gene, was considerably higher than that by E. coli BL21 (DE3)/pE7MAN. Almost mannanase produced by the recombinant E. coli carrying pENS7 at growth temperature of $37^{\circ}C$ existed as inactive enzyme of insoluble form. Growth at temperature below $31^{\circ}C$ increased the soluble fraction of mannanase having catalytic activity in the recombinant E. coli cells. The highest productivity of active mannanase was observed in cell-free extract of the recombinant E. coli grown at growth temperature ranging from $25^{\circ}C$ to $28^{\circ}C$, while mannanase activity per soluble protein of the cell-free extract was highest in the cells grown at $^31{\circ}C$.

• Korean Journal of Microbiology
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• v.46 no.2
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• pp.207-212
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• 2010

A bacterium producing the extracellular mannanase was isolated from Korean soybean paste. The isolate WL-8 has been identified as Bacillus subtilis on the basis on its 16S rRNA sequence, morphology and biochemical properties. The mannanase productivity of strain WL-8 was increased in LB broth by addition of wheat bran. The maximum mannanase productivity was reached to approximately 20 U/ml in LB medium supplemented with 6% wheat bran. A gene encoding the mannanase of WL-8 was cloned into Escherichia coli and its nucleotide sequence was subsequently determined. The mannanase gene consisted of 1,086 nucleotides encoding a polypeptide of 362 amino acid residues. The deduced amino acid sequence was highly homologous with those of several mannanases from B. subtilis belonging to GH family 26. Reaction temperature and pH profiles were investigated using the culture filtrate and cell-free extract of the recombinant E. coli carrying a WL-8 mannanase gene, respectively. Optimal conditions for the two fractions occurred at pH 5.5 and $60^{\circ}C$. The cell-free extract showed higher mannanase activity than the culture filtrate at above $60^{\circ}C$.

• KSBB Journal
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• v.11 no.5
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• pp.565-571
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• 1996

Medium optimization for ${\beta}$-mannanase production by Aspergillus oryzae ATCC 2114 was performed. Effect of carbon source (locust bean gum) concentration on ${\beta}$-mannanase production was investigated. Above 20 g/L locust bean gum, a lag time for ${\beta}$-mannanase production was appeared because high concentration of locust bean gum caused high viscosity which made the mixing of medium poor. As the locust bean gum concentration in the medium increased, ${\beta}$-mannanase activity and cell growth increased proportionally. Effect of various nitrogen sources on ${\beta}$-mannanase production was also studied. (NH4)2SO4 and malt extract were the most effective for ${\beta}$-mannanase production among the inorganic nitrogenous compounds and organic nitrogen nutrients. Inorganic compounds such as KH2SO4, NaCl, Na2CO3, and MgSO4, on ${\beta}$-mannanase production were optimized for ${\beta}$-mannanase production. Locust bean gum of 10 g/L, malt extract of 3 g/L, (NH4)2SO4 of 2 g/L, KH2SO4, of 10 g/L were selected as the optimal medium. Culture in a fermentor by using the optimal medium was carried out. Lag time of ${\beta}$-mannanase production was shorter due to the better mixing of the fermentor. The maximum ${\beta}$- mannanase activity of 9.7 unit/mL and specific ${\beta}$-mannanase activity of 1.9 unit/mg-cell could be obtained at 27 hours and the productivity of ${\beta}$-mannanase was 0.36 unit/mL$.$h.

• Journal of Applied Biological Chemistry
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• v.53 no.2
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• pp.71-76
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• 2010

A gene encoding the mannanase of Bacillus licheniformis WL-12, which had been isolated from Korean soybean paste, was cloned into Escherichia coli and nucleotide sequence of the mannanase gene was subsequently determined. The mannanase gene consisted of 1,080 nucleotides encoding a polypeptide of 360 amino acid residues. The deduced amino acid sequence was identical to that of putative mannanase from B. liceniformis DSM13 belonging to GH family 26. The mannanase was partially purified from cell-free extract of the recombinant Escherichia coli carrying a WL-12 mannanase gene by ammonium sulfate fractionation and DEAE-Sepharose column chromatography. Optimal conditions for the partially purified enzyme occurred at pH 6.0 and $65^{\circ}C$. The enzyme showed higher activity on locust bean gum (LBG) galactomannan and konjac glucomannan than on guar gum galactomannan. The predominant products resulting from the mannanase hydrolysis were mannose, mannobiose and mannotriose for LBG or mannooligosaccharides. The enzyme could hydrolyze mannooligosaccharides larger than mannobiose.

• Journal of Animal Science and Technology
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• v.51 no.5
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• pp.427-432
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• 2009

This study was undertaken to screen a high xylanase and mannanase producing microbes. In the first experiment, screening was undertaken against 50 samples of microorganisms having xylanase and mannanase activities from soil and fallen leaves. The screening process has focused on picking out fungi having high xylanase and mannanase activities under the solid-state fermentation. The xylanase and mannanase activities of 6 screened microbes were 0.9~1.6 unit/mL and 0.2~0.4 unit/mL, respectively, under the submerged fermentation condition. However, under the solid-state fermentation, xylanase and mannanase activities were 103.7~220.0 unit/g and 20.1~40.3 unit/g, respectively. Finally one microbe (E-3) was selected and its xylanase and mannanase activities were 197.3 unit/g and 39.9 unit/g, respectively. The morphological and molecular biological classification of E-3 showed 99% homology with the Aspergillus niger.

• Microbiology and Biotechnology Letters
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• v.37 no.4
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• pp.405-407
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• 2009

A Bacillus subtilis mannanase gene was subcloned into an Escherichia coli- Corynebacterium lactofermentum shuttle vector pHE83, and the resultant plasmid pHE83M was transferred into an endogenous plasmid-free strain of C. lactofermentum. Mannanase produced by the recombinant C. lactofermentum (pHE83M) was secreted extracellulary at the level of 86%, and the remaining activity of mannanase was detected in the cell-free extract. The maximum mannanase productivity of the recombinant strain reached 8.1 unit/mL in the culture filtrate of LB medium. According to the zymogram of mannanase on SDS-PAGE, it was found that the mannanase produced by the recombinant C. lactofermentum could be maintained stably with a migration identical to the mannanase produced by the parental strain, B. subtilis WL-3.

• Korean Journal of Microbiology
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• v.50 no.2
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• pp.158-163
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• 2014

In the acidic LB plate, a bacterial strain was isolated from homemade soybean paste as a producer of the extracellular mannanase. The isolate YB-1402, which was a Gram-positive rod-shaped bacterium with spore, has been identified as Bacillus amyloliquefaciens on the basis of its 16S rDNA sequence and biochemical properties. Maximum mannanase productivity of the isolate YB-1402 was reached approximately 150 U/ml in LB broth supplemented with konjac (3.0%). The molecular mass of YB-1402 mannanase was estimated to approximately 38.0 kDa by zymogram of the culture filtrate on SDS-PAGE. The mannanase of culture filtrate was the most active at $55^{\circ}C$ and pH 5.5. The mannanase activity was completely maintained after pre-incubation at pH 3.0 to 10.0 for 1 h. The predominant products resulting from the mannanase hydrolysis were mannose, mannobiose and mannotriose for LBG or mannooligosaccharides. The enzyme could hydrolyze mannooligosaccharides larger than mannobiose.

• Microbiology and Biotechnology Letters
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• v.25 no.2
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• pp.212-217
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• 1997

A strain of Bacillus sp. WS-14 was isolated from soil. Medium optimization for ${\beta}-mannanase$ production by Bacillus sp. WS-14 was performed. Effect of various carbon sources on ${\beta}-mannanase$ production was investigated and locust bean gum was the most effective for ${\beta}-mannanase$ production. ${\beta}-mannanase$ activity and cell growth increased with increasing the concentration of locust bean gum, however, the amounts were not significant. Among nitrogen sources, soytone was the most effective for ${\beta}-mannanase$ production. Inorganic compounds such as $KH_2PO_4,\;NaCl\;Na_2CO_3\;and\;MgSO_4{\cdot}7H_2O\;on\;{\beta}-mannanase$ production were optimized for ${\beta}-mannanase$ production. Locust bean gum of 10.0 g/l, soytone of 5.0 g/l, $KH_2PO_4$ of 2.0 g/l, NaCl of 10.0 g/l, $MgSO_4{\cdot}7H_2O\;of\;0.2\;g/l,\;Na_2CO_3$, of 2.0 g/l were selected as optimum content. Production of ${\beta}-mannanase$ by using the optimum medium was carried out. The maximum ${\beta}-mannanase$ activity of 20.8 unit/ml could be obtained after 14 h fermentation which corresponed to the productivity of ${\beta}-mannanase$ of 1.48 unit/ml-h.

• Microbiology and Biotechnology Letters
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• v.30 no.3
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• pp.247-252
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• 2002

A bacterium producing the extracellular mannanase was isolated from Korean formented food and has been identified as a member of the genus Bacillus from the result of the phylogenic analysis based on partial 165 rRNA sequences. The isolate, named Bacillus sp. WL-3, was shown to be similar to B. subtilis strain on the basis of its biochemical properties. The mannanase of culture supematant was the most active at $55^{\circ}C$ and pH 6.0. The additional carbohydrates including u-cellulose, avicel, oat spelt xylan, guar gum and locust bean gum (LBG) increased the mannanase productivity. Especially, the maximum mannanase productivity was reached 65.5 U/ml in LB medium supplemented with 0.5% (w/v) LBG, which was 131-folds more than that in LB medium. It was sug-gested that the increase of mannanase production was owing to induction of mannanase biosynthesis by LBG hydrolysates transported following initial hydrolysis by extracellular mannanase during the cell growth. The molec-ular weight of WL-3 mannanase was estimated to approximately 38.0 kDa by zymogram on SDS-PAGE.

• Microbiology and Biotechnology Letters
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• v.43 no.3
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• pp.204-211
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• 2015

A bacterial strain capable of hydrolyzing xylan and locust bean gum (LBG) was isolated from the Saemangeum tideland of Korea. Based on the biochemical properties and the 16S rRNA gene sequence, the isolate YB-30 was identified as Bacillus subtilis. Xylanase productivity was increased effectively when B. subtilis YB-30 was grown in the presence of wheat bran, while mannanase productivity was increased drastically when grown in the presence of konjac or LBG. Particularly, maximum mannanase and xylanase activities were detected in the culture filtrate of media containing 3.5% konjac and 1% wheat bran. Both enzyme productivities reached maximum levels in the stationary growth phase. The culture filtrate exhibited the highest activity at 60℃ and pH 6.0 for mannanase and at 55℃ and pH 5.5 for xylanase, respectively. Both enzymes were not stable at high temperatures and xylanase was less stable than mannanase. In addition, wheat bran was hydrolyzed to liberate reducing sugar to a greater extent than rice bran by the culture filtrate because the wheat bran contained more arabinoxylan than the rice bran. Hence, xylanase and mannanase produced by B. subtilis YB-30 have a potential use as feed additive enzymes.