• Korean Journal of Microbiology
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• v.37 no.4
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• pp.253-258
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• 2001

A Bacillus circulans KCTC3004 $\beta$-1,3-glucanase gene contained in a recombinant plasmid pLM460 derived from subcloning the original recombinant plasmid pLM530 was trasferred into a new shuttle vector plasmid pLMS1180 by ligating linearized DNAs of pLM460 and pUB110. B. subtilis RM125 and B. megaterium ATCC14945 transformed with pLMS1180 produced the $\beta$-1,3-glucanase substantially. Most of the enzyme was produced during the exponential growth period. The maxium activities of the $\beta$-1,3-glucanase produced by the Bacillus transformants were compared with that of the B. circulans gene donor strain. The B. subtilis RM125 (pLM1180) enzyme showed the activity 14 times higher than that of the gene donor cells, followed by the B. megaterium ATCC14945 (pLMS 1180) enzyme with activity 5 times higher than that of the gene donor cells. While E. coli secreted about 7% of the produced enzyme, B. subtilis excreted the enzyme into the medium wholly and B. megaterium about 97% of the total product. The SDS-PAGE of this enzyme produced in E. coli (pLMS1180), B subtilis (pLMS1180) or B. megaterium (pLMS1180) indicated a molecular weight of 38,000. The enzymes overproduced in three different host cells hydrolyzed laminarin to produce mainly laminaribiose, laminaritriose, and laminarioligosaccharides. The plasmid pLMS1180 was stable in B. megaterium, E. coli, but was unstable in B. subtilis.

• Korean Journal of Microbiology
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• v.42 no.1
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• pp.67-72
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• 2006

The nucleotide sequence of the cloned cellulolytic xylanase gene (bglBC2) from B. circulans ATCC21367 was determined. bglBC2 consists of an 1,224 bp open reading frame (ORF) coding for a polypeptide of 407 amino acids with a deduced molecular weight of 45 kDa. The Shine-Dalgarno (SD) sequence (5'-AAAGGAG-3') was found 9 bp upstream of the initiation codon, ATG. A promoter region corresponding closely to the B. subtilis consensus sequence (-35: TTGACA,-10: TATAAT) was detected, the putative -35 and -10 sequences of which were TTTACA and TATACT, respectively. The deduced amino acid sequence of the cellulolytic xylanase showed 97% homology with that of the alkaline $endo-\beta-1,4-glucanase$ from B. circulans KSM-N257, 75% homology with that of the $endo-\beta-1,3-1,4-glucanase$ from B. circulans WL-12, and 45% homology with that of the $endo-\beta-1,4-glucanase$ (cellulase) from Bacillus sp. KSM-330. The bglBC2 sequence was deposited in Gen-Bank under the accession number AY269256.

• Journal of Microbiology
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• v.33 no.4
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• pp.295-301
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• 1995

An antifungal Pseudomonas stutzeri YPL-1 produced extracellular chitinase and .betha.-1, 3-glucanase that were key enzymes in the decomposition of fungal hyphal walls. These lytic extracellular enzymes markedly inhibited mycelial growth of the phytopathogenic fungus Fusarium solani. A chitinase from P. stutzeri YPL-1 inhibited fungal mycelial growth by 87%, whereas a .betha.-1, 3-glucanase from the bacterium inhibited growth by 53%. Furthermore, co-operative action of the enzymes synergistically inhibited 95% of the fungal growth. The lytic enzymes caused absnormal swelling and retreating on the fungal hyphal walls in a dual cultures. Scanning electron microscopy clearly showed hyphal degradation of F. solani in the regions interacting with P. stutzeri YPL-1. In an in vivo pot test, P. stutzeri YPL-1 proved to have biocontrol ability as a powerful agent in controlling plant disease. Planting of kidney bean (Phaseolus vulgaris L.) seedlings with the bacterial suspension in F. solani-infested soil significantly suppressed the development of fusarial root-rot. The characteristics of a crude preparation of .betha.-1, 3-glucanase produced from P. stutzeri YPL-1 were investigated. The bacterium detected after 2 hr of incubation. The enzyme had optimum temperature and pH of 40.deg.C and pH 5.5, respectively. The enzyme was stable in the pH range of 4.5 to 7.0 and at temperatures below 40.deg.C, with a half-life of 40 min at 60.deg.C.

• Journal of Plant Biotechnology
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• v.43 no.4
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• pp.450-456
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• 2016

Rhizoctonia leaf blight (large patch) has become a serious problem in Korean lawn grass, which is extremely hard to treat and develops mostly from the roots of lawn grass to wither it away. Rhizoctonia leaf blight (large patch) is caused by Rhizoctonia solani AG2-2 (IV). To develop zoysia japonica with strong disease tolerance against this pathogenic bacterium, ${\beta}-1,3-glucanase$ was cloned from zoysia japonica, which is one of the PR-Proteins known to play a critical role in plant defense reaction. ${\beta}-1,3-glucanase$ is known to be generated within the cells when plant tissues have a hypersensitive reaction due to virus or bacterium infection and secreted outside the cells to play mainly the function of resistance against pathogenic bacteria in the space between the cells. This study utilized the commonly preserved part in the sequence of corn, wheat, barley, and rice which had been researched for their disease tolerance among the ${\beta}-1,3-glucanase$ monocotyledonous plants. Based on the part, degenerate PCR was performed to find out the sequence and full-length cDNA was cloned. E.coli over-expression was conducted in this study to mass purify target protein and implement in vitro activation measurement and antibacterial test. In addition, to interpret the functions of ZjGlu1 gene, each gene-incorporating plant transformation vectors were produced to make lawn grass transformant. Based on ZjGlu1 protein, antibacterial activity test was conducted on 9 strains. As a result, R. cerealis, F. culmorum, R.solani AG-1 (1B), and T. atroviride were found to have antibacterial activity. The gene-specific expression amount in each organ showed no huge difference in the organs based upon the transformant and against 18s gene expression amount.

• Microbiology and Biotechnology Letters
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• v.16 no.2
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• pp.79-84
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• 1988

Dye materials and cross linking agents were used for the determination of $\beta$-glucanase activities. The objective of this study was to prepare the blue coloured substrates which are sensitive, specific and simple for the determination of $\beta$-glucanase in malt and Bacillus subtilis K-4-3 enzymes. This method is based on the principle of measuring colorimetrically the split product of coloured and cross linked substrate. The best coupling of dye stuff of $\beta$-glucan was cibacron blue 3G-A and the colour released can suitably be measured at 623nm. Optimal concentration of dye and cross linking agents was 1.5g and 1.25$m\ell$ under 0.1N NaOH. The sensitivity comparison proved that the stained $\beta$-glucan method is much more sensitive than the DNS method to determine reducing sugar released by the enzyme.

• Mycobiology
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• v.45 no.4
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• pp.385-391
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• 2017

The ability of Bacillus subtilis, strain ALICA to produce three mycolytic enzymes (chitinase, ${\beta}$-1,3-glucanase, and protease), was carried out by the chemical standard methods. Bacillus subtilis ALICA was screened based on their antifungal activity in dual plate assay and cell-free culture filtrate (25%) against five different phytopathogenic fungi Alternaria alternata, Macrophomina sp., Colletotrichum gloeosporioides, Botrytis cinerea, and Sclerotium rolfesii. The B. subtilis ALICA detected positive for chitinase, ${\beta}$-1,3-glucanase and protease enzymes. Fungal growth inhibition by both strain ALICA and its cell-free culture filtrate ranged from 51.36% to 86.3% and 38.43% to 68.6%, respectively. Moreover, hyphal morphological changes like damage, broken, swelling, distortions abnormal morphology were observed. Genes expression of protease, ${\beta}$-1,3-glucanase, and lipopeptides (subtilosin and subtilisin) were confirmed their presence in the supernatant of strain ALICA. Our findings indicated that strain ALICA provided a broad spectrum of antifungal activities against various phytopathogenic fungi and may be a potential effective alternative to chemical fungicides.

• Journal of Microbiology and Biotechnology
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• v.31 no.10
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• pp.1446-1454
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• 2021

Herein, we cloned and expressed an endo-β-1,4-glucanase gene (celA1805) from Bacillus subtilis B111 in Escherichia coli. The recombinant celA1805 contains a glycosyl hydrolase (GH) family 8 domain and shared 76.8% identity with endo-1,4-β-glucanase from Bacillus sp. KSM-330. Results showed that the optimal pH and temperature of celA1805 were 6.0 and 50℃, respectively, and it was stable at pH 3-9 and temperature ≤50℃. Metal ions slightly affected enzyme activity, but chemical agents generally inhibited enzyme activity. Moreover, celA1805 showed a wide substrate specificity to CMC, barley β-glucan, lichenin, chitosan, PASC and avicel. The K_m and V_max values of celA1805 were 1.78 mg/ml and 50.09 µmol/min/mg. When incubated with cellooligosaccharides ranging from cellotriose to cellopentose, celA1805 mainly hydrolyzed cellotetrose (G4) and cellopentose (G5) to cellose (G2) and cellotriose (G3), but hardly hydrolyzed cellotriose. The concentrations of reducing sugars saccharified by celA1805 from wheat straw, rape straw, rice straw, peanut straw, and corn straw were increased by 0.21, 0.51, 0.26, 0.36, and 0.66 mg/ml, respectively. The results obtained in this study suggest potential applications of celA1805 in biomass saccharification.

• Journal of Microbiology and Biotechnology
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• v.22 no.3
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• pp.407-415
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• 2012

An endophytic bacterium, Bacillus thuringiensis GS1, was isolated from bracken (Pteridium aquilinum) and found to have maximal production of chitinase (4.3 units/ml) at 5 days after culture. This study investigated the ability of B. thuringiensis GS1 to induce resistance to Rhizoctonia solani KACC 40111 (RS) in cucumber plants. Chitinase activity was greatest in RS-treated plants at 4 days. ${\beta}$-1,3-Glucanase activity was highest in GS1-treated plants at 5 days. Guaiacol peroxidase (GPOD) activity increased continuously in all treated plants for 5 days. Ascorbate peroxidase (APX) activity in RS-treated plants was increased 1.5-fold compared with the control at 4 days. Polyphenol oxidase (PPO) activity in RS-treated plants was increased 1.5-fold compared with the control at 3 days. At 5 days after treatment, activity staining revealed three bands with chitinase activity (Ch1, Ch2, and Ch3) on SDS-PAGE of cucumber plants treated with GS1+RS, whereas only one band was observed for RS-treated plants (Ch2). One GPOD isozyme (Gp1) was also observed in response to treatment with RS and GS1+RS at 4 days. One APX band (Ap2) was present on the native-PAGE gel of the control, and GS1- and GS1+RS-treated plants at 1 day. PPO bands (Po1 and Po2) from RS- and GS1+RS-treated plants were stronger than in the control and GS1-treated plants upon native-PAGE at 5 days. Taken together, these results indicate that the induction of PR proteins and defense-related enzymes by B. thuringiensis GS1 might have suppressed the damping-off caused by R. solani KACC 40111 in cucumber plants.

• Korean Journal of Microbiology
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• v.54 no.4
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• pp.354-361
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• 2018

This study was carried out to isolate soil bacteria with antimicrobial activity and evaluate antimicrobial substances produced by isolated bacteria. Among many isolates Bacillus subtilis DS660 and Paenibacillus polymyxa DS842 showed high antimicrobial activities against 6 species of microbial residents on human skin and 3 species of pathogenic bacteria. DS660 and DS842 showed 15.3~26.8 and 11.3~27.5 mm of inhibition zone diameter, respectively on nutrient agar medium against most target bacteria and fungi. DS660 and DS842 produced $57{\pm}8$ and $170{\pm}15{\mu}mol/ml$ of siderophore, respectively as an antimicrobial substance. Analysis of ethyl acetate extract of culture supernatants of DS660 and DS842 suggested production of glycolipid biosurfactant which reduced surface tension of culture supernatant of DS660 and DS842 from 60.0 to 40.3 and 30.3 mN/m, respectively. DS660 and DS842 also showed $169.2{\pm}9.9$ and $357.2{\pm}13.7nmol/min/mg$ protein of ${\beta}-1,3$-glucanase activity, respectively, and hydrolyzed cell wall components of 3 bacterial species. These results suggest that B. subtilis DS660 and P. polymyxa DS842 may be utilized as an environment-friendly biocontrol agent against some skin microbes and pathogenic bacteria.

• Asian-Australasian Journal of Animal Sciences
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• v.26 no.2
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• pp.253-259
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• 2013

Paddy rice is rarely used as a feed because of its high fiber content. In this study, two experiments were conducted to study the effects of supplementing an enzyme complex consisting of xylanase, beta-glucanase and cellulase, to paddy-based diets on the performance and nutrient digestibility in meat-type ducks. In the both experiments, meat-type ducks (Cherry Valley) were randomly assigned to four treatments. Treatment 1 was a basal diet of corn-soybean; treatment 2 was a basal diet of corn-paddy-soybean; treatment 3, had enzyme complex added to the corn-paddy-soybean basal diet at levels of 0.5 g/kg diet; and treatment 4, had enzyme complex added to the corn-paddy-soybean diet at levels of 1.0 g/kg diet. The results showed that the enzyme complex increased the ADG, and decreased the ADFI and F/G significantly (p<0.05) in the ducks, and the ADFI for the ducks fed the corn-paddy-soybean diet showed no difference compared to the ducks fed corn-soybean diets at all stages of the experiment (p<0.05). When corn was partially replaced by paddy, the digestibility of CP and NDF was decreased and increased, respectively (p<0.05), and the level of enzyme complex had a significant effect on both CP and NDF digestibility (p<0.05). As for the AME, addition of enzyme complex increased it significantly (p<0.05), but both diet types and levels of enzyme complex had no effect (p>0.05). The outcome of this research indicates that the application of enzyme complex made up of xylanase, beta-glucanase, and cellulase, in the corn-paddy-soybean diet, can improve performance and nutrition digestibility in meat-type ducks.