• Microbiology and Biotechnology Letters
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• v.17 no.6
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• pp.545-551
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• 1989

Thermostable alkaline protease of alkalophilic Bacillus sp. No. 8-16 has been purified, and the properties of the enzyme investigated. The characteristic point of the organism used is especially good growth in alkaline and thermal condition. The alkaline protease of the strain No. 8-16 was purified from crude enzyme by acetone precipitation, CM-cellulose ion exchange chromatography, Sephadex G-100 and Sephadex G-75 gel filtration. Through the series of chromatograpies, the enzyme was purified to homogeneity with specific activity of 37 fold higher than that of the crude broth. Characteristics of the purified enzyme were as follow; $K_m$ value for the enzyme was 1.3 mg/ml, the alkaline protease showed a maximal activity at 7$0^{\circ}C$ and from the pH 6.0 through 12.0, and stable for 1 hr. at 6$0^{\circ}C$. The moleclar weight of the enzyme was estimated to be 33,000 by Sephadex G-100 gel filtration. The activity of the alkaline protease was inhibited by iodoacetic acid and Ag$^+$, Hg$^+$, PMSF (phenylmethylsulfonyl fluoride), and activated by $Ca^{2+}$ and Mn$^{2+}$.

• Journal of Microbiology and Biotechnology
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• v.7 no.3
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• pp.212-214
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• 1997

A new isolate, Bacillus sp. ALK-7 can synthesize ethylene from l-aminocyclopropane-l-carboxylic acid (ACC) as well as from methionine. The ACC has only been recognized as a key intermediate found in the metabolic pathway leading to ethylene formation in various plants. The efficiency of ethylene formation from the ACC by Bacillus sp. ALK-7 was about 2 times as high as that from the methionine. The reaction from ACC to ethylene formation was also shown to be mediated by the cell-free extracts of Bacillus sp. ALK-7.

• Microbiology and Biotechnology Letters
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• v.26 no.3
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• pp.195-199
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• 1998

AH alkalophilic Bacillus SP. AIk-7, isolated from soil, produced ethylene. The characteristics of this microorganism is the ability to grow well under the alkaline condition, at pH 10.3. This strain is similar to Bacillus alkalophilus in terms of morphological, physiological and biological characteristics. In observation of relationship of cell growth and ethylene production according to incubation times, the ethylene synthesis mostly occur from the late exponential phase to the death phase of growth. The purpose of this paper is to study the effects of various substrates on the biosynthesis of ethylene in the intact cell and the cell-free system by the Bacillus sp. AIk-7. In both intact cell and cell-free extract, optimum conditions for ethylene production was achieved at pH 10.3 and 3$0^{\circ}C$. Ethylene was effectively produced from L-Met and 1-aminocyclopropane-1-carboxylic acid (ACC). In this case, ACC as the substrate on ethylene production were two fold higher than L-met at each concentration of substrates. On the other hand, the cell-free ethylene-forming system was used as a tool for the elucidation of the biochemical reaction involved in the formation of ethylene by Bacillus sp. AIk-7. Ethylene production in the cell-free system required the presence of manganese and cobalt ion to be stimulated a little. The result obtained in this work suggests that L-met and ACC may be a precursor more directly related to bacterial ethylene production than any other substrates tested.

• Microbiology and Biotechnology Letters
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• v.25 no.1
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• pp.30-36
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• 1997

To genetically breed powerful multifunctional antagonistic bacteria, the urease gene of alkalophilic Bacillus pasteurii was transferred into Bacillus subtilis YB-70 which had been selected as a powerful biocontrol agent against root-rotting fungus Fusarium solani. Urease gene was inserted into the HindIII site of pGB215-110 and designated pGU266. The plasmid pGU266 containing urease gene was introduced into the B. subtilis YB-70 by alkali cation transformation system and the urease gene was very stably expressed in the transformant of B. subtilis YB-70(pGU266). The optimal conditions for the transfomation were also evaluated. From the in vitro antibiosis tests against F. solani, the antifungal activity of B. subtilis YB-70 containing urease gene was much efficient than that of the non-transformed strain. Genetic improvement of B. subtilis YB-70 by transfer of urease gene for the efficient control seemed to be responsible for enhanced plant growth and biocontrol efficacy by combining its astibiotic action and ammonia producing ability.

• The Journal of Natural Sciences
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• v.5 no.2
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• pp.13-17
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• 1992

Regulation of $\beta$-galactosidase biosynthesis was studied with alkalophilic, thermophilic Bacillus sp. TA-11. Biosynthesis of the enzyme was effectively induced by lactose and some low level by isoprophyl-$\beta$-D-thiogalactopyranoside(IPTG). When 30mM glucose was added at the different intervals to the culture that had been in contact with lactose, the different levels of the enzyme synthesis were observed. So, this suggests that glucose interfered with the entry of the lactose into the cells.The glucose inhibitory effect was not relieved by adding cAMP to the culture.

• Microbiology and Biotechnology Letters
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• v.17 no.5
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• pp.436-439
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• 1989

The chromosomal DNA fragments of thermophilic alkalophilic Bacillus sp, K-17, a potent xylanhydrolyzing bacterium, were ligated to a vector plasmid pBR322 and transformed into Escherichia coli HB101. The plasmid pAX278, isolated from a transformant forming yellow color on the LB agar plate containing 1 mM p-nitrophenyl- $\beta$-xylopyranoside, was found to enable the transformants to produce p-xylosidase. The 5.0 kilobase insert of pAX278 had single sites for EcoRI, PstI, XbaI, and PvuII, and 2 sites for BglII. Biotinylated pAX218 was hybridized to 0.9 kb as well as 5.0 kb fragment from Bacillus sp. K-17 DNA on nitrocellulose filter. pGX718 was constructed by inserting the 5.0 kb HindIII fragment of pGX278 at the HindIII site of pGR71, E. coli and B. subtilis shuttle vector. The enzymatic properties of $\beta$-xylosidase from E. coli HB101 carrying recombinant plasmid were the same those of $\beta$-xylosidase from Bacillus sp. K-17.

• Microbiology and Biotechnology Letters
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• v.23 no.6
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• pp.710-716
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• 1995

$\beta $-1, 4-D-arabinogalactanase isolated from alkalophilic Bacillus sp. HJ-12, approximate Mw 42 kDa, was generally stable in the range of pH 6-10 and below 50$\circ$C and its highest activity was observed at 60$\circ$C with pH 7-9. The isolated $\beta $-1, 4-D-arabinogalactanase specifically hydrolyzed $\beta $-1, 4-galactosyl linkage that is the major structure of soybean arabinogalactan (SAG) but not $\beta $-1, 3-galactosyl linkage of the other polysaccharides. K. was estimated as 0.67 mg/ml by the method of Hanes-Woolf plot. No metals and chemical reagents inhibited the enzyme activity but urea did. The active site of this enzyme assumed to be tryptophan residue. The hydrolysis products from SAG, assayed by gel chromatography, TLC and HPLC, were predominantly galactotetraose (Gal$_{4}$) and triose (Gal$_{3}$) with a small portion. $\beta $-1, 4-D-arabinogalactanase hydrolyzed ONPG as well as SAG, and the degree of hydrolysis of SAG was 15% which is lower than that by the other $\beta $-1, 4-galactanases from different sources. SAG treated with this enzyme resulted in the reduction of specific viscosity up to 70%.

• KSBB Journal
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• v.9 no.4
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• pp.400-405
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• 1994

The conditions for ${\beta}$-galactosidase production from alkalophilic, thermophilic Bacillus sp. TA-11 were investigated. The maximal enzyme production was obtained when the strain was cultured at $50^{\circ}C$ for 5 days with fed-batch culture in the optimal medium containing 1.5% lactose, 0.6% yeast extract 0.15% $K_2HP0_4$and initial pH 9.5, and then final enzyme activity under the above conditions was 5200 unit/ml of cell free extract.

• Microbiology and Biotechnology Letters
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• v.17 no.4
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• pp.283-288
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• 1989

To reduce the size of 5.0kb HindIII fragment containing $\beta$-xylosidase gene, the 5.0kb insert of pAX278 which was previously cloned was reduced by various deletions and thus 1.4kb EcoRI-Xbal fragment was subcloned into pUC19, and the recombinant plasmid was named pAK208. The $\beta$-xylosidase acnivity of E. coli harboring pAK208 was higher about 1.3times than that of pAX278. For the improvement of $\beta$-xylosidase activity, we cloned and expressed the $\beta$-xylosidase gene in E. coli using vector pKK223-3 containing a potent tac-promoter, and enzyme activity of the transformant harboring pKHR212 was increased about 3.3 and 1.8 times than that of E. coli(pAX278) and Bacillus sp. K-17, respectively. To obtain better expression of $\beta$-xylosidase gene, the whole 5.0kb HineIII fragment was recloned into pC194, and the Bacillus sp. K-17 transformant harbor-ing the recombinant plasmid pCX174 showed higher activity than that of the E. coli (pAX278) and Bacillus sp. K-17, respectively. The characteristics of enzyme purified from transformants were consistent with those front alkalophilic Bacillus sp, K-17.

• Microbiology and Biotechnology Letters
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• v.44 no.3
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• pp.333-340
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• 2016

An alkaline protease was purified and characterized from an alkalophilic microorganism, Bacillus sp. DK1122, isolated from soil in central Korea. The optimum temperature and pH for the growth of the producer strain were 40℃ and pH 9.0, respectively. The protease was produced aerobically at 40℃ after 24 h incubation in modified Horikoshi I medium (pH 9.0) containing 0.5% (w/v) glucose, 0.8% (w/v) yeast extract, 0.5% (w/v) polypeptone, 0.1% (w/v) K₂HPO₄, 0.02% (w/v) MgSO₄·7H₂O, 1% (w/v) Na₂CO₃, and 3% (w/v) NaCl. The alkaline protease was purified by 70% ammonium sulfate precipitation of the culture supernatant of Bacillus sp. DK1122, followed by CM-Sepharose chromatography. The molecular weight of the enzyme was estimated to be 27 kDa on the basis of SDS-PAGE. The optimum temperature and pH for the protease activity were 60℃ and pH 9.0, respectively. Addition of CaCl₂ increased the thermal stability of the purified protease, where 90% of protease activity was retained at 60℃ for up to 3 h. Consequently, it is expected that the alkaline protease from this study, exhibiting stability at pH 7–9 and 60℃, may be promising for application in the food and detergent industries.