• 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.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.

• KSBB Journal
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• v.14 no.6
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• pp.718-723
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• 1999

In order to treat of alkaline dye-processing wastewater, alkalophilic strains biodegrading azo dye, Acid red 1, is isolated from natural system, and optimal culture conditions are examined using response surface analysis, statistical analysis system program. 15 different species which grow in alkaline culture media are isolated from the effluent and river soil discharged from wastewater treatment plant in dye industrial complex. One strain which has the best decolorization efficiency is chosen, and named as AR-1. The result of the examination of carbon, nitrogen and phosphorus sources which have influence on growth and decolorization reveals that optimum carbon, nitrogen and phosphorus sources are 1.0% fructose, 1.0% polypeptone, 1.0% yeast extract and 0.5% $K_2HPO_4$, respectively. In order to optimize of biodegradation conditions of dye by response surface analysis, the characteristics of decolorization and cell growth according to culture temperature and time are monitered. The result shows that the one is optimum 34.77$^{\circ}C$ for 12.97 hours; the other at 34.73$^{\circ}C$ for 12.96 hours. While, optimal conditions of culture that satisfy both cell growth and decolorization are the temperatures from 32.86$^{\circ}C$ to 36.36$^{\circ}C$ and the period of 10.96 to 15.75 hours, respectively.

• 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.16 no.4
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• pp.275-281
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• 1988

To develop cellulase and xylanase-producing strain by protoplast fusion, alkalophilic Bacillus sp. F204 and K17 were treated with NTG(N-methyl-N'-nitro-N-nitrosoguanidine) and isolated anti-biotics resistant strains of S20 (Km$^r$ , Cm$^r$) and G70 (Str$^r$). The frequency of protoplast formation was about 95% when cells of mid-log phase were treated with 200$\mu\textrm{g}$/ml Iysozyme at 37$^{\circ}C$ for 30-45 minutes. Under addition of 0.4-0.5M sodium succinate, 0.5% casamino acid, 1.5% polyvinylpyrrolidone, 25mM MgC1$_2$ and 50mM CaC1$_2$ to the regeneration medium, the regeneration frequency of Bacillus sp. F204 and K17 was 24.9% and 26.2%, respectively. The fusion frequency was 6.6$\times$10$^{-6}$ in the presence of 30% polyethylene glycol 6000 containing 50mM $Ca^{++}$ at 45$^{\circ}C$ for 5 minutes. Cellulase complex and xylanase activities of fusant were compared with parental strains.

• 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.21 no.5
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• pp.453-460
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• 1993

The extracellular bacteriolytic enzyme from alkalophilic Bacillus sp. YJ-451 was endopeptidase which hydrolyzes the peptide bond at the amino group of D-glutamic acid in the peptidoglycan. Protoplast transfomation system of B. subtilis by the lytic enzyme that differs, in mechanisms, from lysozyme which was used to transformation of B. subtilis was investigated. High protoplast yield was obtained from cells cultured in PAB at the late logarithmic growth phase.

• Journal of the East Asian Society of Dietary Life
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• v.10 no.5
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• pp.439-444
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• 2000

호알칼리성 방선균 Streptomyes sp. B-2를 Glucose Isomerse 생성을 위해 토양에서 분리했다. Glucose Isomerase(G.I)는 high fructose glucose syrup과 fructose의 생산을 위해서 식품 공업에서 아주 중요시되고 있는 효소이다. 호알칼리성 방선균 Streptomyces sp. B-2가 생성하는 glucose isomerase(G.I.)를 정제하였다. G.I.는 (NH$_4$)$_2$So$_4$분획, DEAE-cellulose, Sephadex G-200 chromatography하여 순수 분리 하였다. 순수분리된 G.I.는 electrophoresis에 의해 확인을 했다. SDS-acrylamide gel electrophoresis에 의해 정제된 효소는 single band를 보여주었다.

• Korean Journal of Microbiology
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• v.47 no.3
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• pp.194-199
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• 2011

An alkalophilic bacterium producing alkaline protease was isolated from waste water and solar saltern sample and identified as Bacillus pseudofirmus HS-54 based on morphological, biochemical characteristics as well as 16S-rRNA gene sequencing. The HS-54 protease was purified to homogeneity using ammonium sulfate precipitation, DEAE cellulose column chromatography, and sephadex G-100 gel filtration with a 4.0 purification fold. The molecular mass of the purified enzyme was estimated by SDS-PAGE to be 27 kDa. The optimal pH and temperature for the purified protease activity were 10.0 and $50^{\circ}C$, respectively. The purified enzyme was relatively stable at the pH range of 6.0-11.0 and at the temperature below $50^{\circ}C$. This enzyme was activated by $Ca^{2+}$ and $Mg^{2+}$ and inhibited by $Hg^{2+}$, $Cu^{2+}$, $Zn^{2+}$, $Al^{3+}$, $Ag^{2+}$. And this enzyme was strongly inhibited by PMSF, suggesting that it belongs to the serine protease superfamily.

• Journal of Microbiology and Biotechnology
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• v.14 no.6
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• pp.1142-1149
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• 2004

Streptococcus mutans has the capacity of inducing dental caries. Thus, to develop a novel way of preventing dental caries, a cell wall hydrolase-producing strain was isolated and its characteristics were investigated. Among 200 alkalophilic strains isolated from soil, 8 strains exhibited lytic activities against Streptococcus mutans. However, strain YU5215 with the highest cell wall hydrolase activity was selected for further study. Strain YU5215 was identified as a novel strain of Bacillus based on analyzing its 16S rDNA sequence and Bergey's Manual of Systematic Bacteriology, and thus designated as Bacillus mutanolyticus YU5215. The optimal conditions for the production of the cell wall hydrolase from Bacillus mutanolyticus YU5215 consisted of glucose ($0.8\%$), yeast extract ($1.2\%$), polypeptone ($0.5\%$), $K_{2}HPO_{4}\;(0.1\%$), $MgSO_{4}{\cdot}7H_{2}O$ ($0.02\%$), and $Na_{2}CO_{3}\;(1.0\%$) at pH 10.0. Bacillus mutanolyticus YU5215 was cultured at 30^{circ}C for 72 h to produce the cell wall hydrolase, which was then purified by acetone precipitation and CM-agarose column chromatography. The molecular weight of the lytic enzyme was determined as 22,700 Da by SDS-PAGE. When the cell wall peptidoglycan of Streptococcus mutans was digested with the lytic enzyme, no increase in the reducing sugars was observed, while the free amino acids increased, indicating that the lytic enzyme had an endopeptidase-like property. The amino terminus of the cell wall peptidoglycan digested by the lytic enzyme was determined as a glutamic acid, while the lytic site of the lytic enzyme in the Streptococcus mutans peptidoglycan was identified as the peptide linkage of L-Ala and D-Glu.