• Applied Biological Chemistry
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• v.46 no.3
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• pp.246-250
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• 2003

In order to isolate antibacterial substances from the rhizome of Zingiber mioga Roscoe, the ethanol extracts was fractionated according to the activity against Bacillus subtilis, B. cereus and Staphylococcus aureus. Three antibacterial substances were isolated and purified by column chromatography and recrystallization. Compounds I and III showed activity against all the tested bacterias and compound II exhibited the activity against B. subtilis and B. cereus S. aureus. Compound I was examined antimicrobial activity against B. subtilis, B. cereus and S. aureus by optical density using Bioscreen C. Compound I showed strong growth inhibition at 10 ppm on B. subtilis and B. cereus for 72 hrs, and at 25 ppm on S. aureus. On the basis of spectrometric studies including $1^H-NMR$, ${13}^C-NMR$, DEPT, IH-lH COSY, HMQC, HMBC and IR, compounds I, II and III were identified as $(E)-8{\beta}(17)-epoxylabd-12-ene-15,16-dial\;(C_{20}H_{30}O_3,\;MW=318)$, galanolactone $(C_{20}H_{30}O_3\;MW=318)$ and galanal A $(C_{20}H_{30}O_3,\;MW=318)$, respectively. These results are the first reports on the isolation of $(E)-8{\beta}(17)-epoxylabd-12-ene-15,16-dial, galanolactone and galanal A from the rhizome of Zingiber mioga.

• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1514-1519
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• 2009

A gene encoding the xylanase of Bacillus subtilis AMX-4 isolated from soil was cloned into Escherichia coli and the gene product was purified from the cell-free extract of the recombinant strain. The gene, designated xylA, consisted of 639 nucleotides encoding a polypeptide of 213 residues. The deduced amino acid sequence was highly homologous to those of xylanases belonging to glycosyl hydrolase family 11. The molecular mass of the purified xylanase was 23 kDa as estimated by SDS-PAGE. The enzyme had a pH optimum of 6.0-7.0 and a temperature optimum of $50-55^{\circ}C$. Xylanase activity was significantly inhibited by 5 mM $Cu^{2+}$ and 5 mM $Mn^{2+}$, and noticeably enhanced by 5 mM $Fe^{2+}$. The enzyme was active on xylans including arabinoxylan, birchwood xylan, and oat spelt xylan, but it did not exhibit activity toward carboxymethylcellulose or p-nitrophenyl-$\beta$-xylopyranoside. The predominant products resulting from xylan and xylooligosaccharide hydrolysis were xylobiose and xylotriose. The enzyme could hydrolyze xylooligosaccharides larger than xylotriose.

• Journal of Microbiology and Biotechnology
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• v.25 no.9
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• pp.1493-1501
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• 2015

A Bacillus species, EMD4, with strong antibacterial activity was isolated from ganjang (soy sauce) and identified as B. subtilis. B. subtilis EMD4 strongly inhibited the growth of B. cereus ATCC14579 and B. thuringiensis ATCC33679. The antibacterial activity was stable at pH 3-9 but inactive at pH 10 and above. The activity was fully retained after 15 min at 80℃ but reduced by 50% after 15 min at 90℃. The activity was completely destroyed by proteinase K and protease treatment, indicating its proteinaceous nature. The bacteriocin (BacEMD4) was partially purified from culture supernatant by ammonium sulfate precipitation, and Q-Sepharose and Sephadex G-50 column chromatographies. The specific activity was increased from 769.2 AU/mg protein to 8,347.8 AU/mg protein and the final yield was 12.6%. The size of BacEMD4 was determined to be 3.5 kDa by Tricine SDS-PAGE. The N-terminal amino acid sequence was similar with that of Subtilosin A. Nucleotide sequencing of the cloned gene confirmed that BacEMD4 was Subtilosin A. BacEMD4 showed bactericidal activity against B. cereus ATCC14579.

• BMB Reports
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• v.32 no.3
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• pp.239-246
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• 1999

A tripeptidase (PepT) was purified to homogeneity from Bacillus subtilis through four sequential chromatographies including DEAE-Sepharose ion exchange, hydroxylapatite, mono-Q FPLC ion exchange, and Superose-12 FPLC gel filtration. The apparent molecular mass of the enzyme was 49,200 Da and 51,400 Da as determined by sodium dodecylsulfatepolyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography, respectively, and the enzyme exists in a monomeric form. The physicochemical properties of the enzyme were as follows: optimum pH at 7.5, optimum temperature at $60^{\circ}C$, and pI at 4.9. The $K_m$ and $V_{max}$ values of the enzyme were 4.3 mM and 2.5 mmol/min/mg, respectively, with MetAla-Ser as substrate. The B. subtilis PepT requires $Co^{2+}$ ion(s) for activation, while it is inactivated by EOTA and 1,10-phenanthroline, suggesting that it is a metalloprotein. The enzyme was not inhibited by any of serine protease, aspartic protease, or leucine aminopeptidase inhibitors. The enzyme showed comparable activities towards four different substrates including Met-Ala-Ser, Leu-Gly-Gly, Leu-Ser-Phe, and Leu-Leu-Tyr. The amino terminal sequence of PepT determined by Edman degradation was found to be MKEEIIERFTTYVXV and turned out to be identical to that of PepT deduced from a cloned B. subtilis pepT.

• Applied Biological Chemistry
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• v.31 no.4
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• pp.356-360
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• 1988

The alkaline protease producing bacteria isolated from soil and identified as Bacillus subtilis. The optimum medium for alkaline protease production from the microorganism was as follows; soluble starch, 1.5% ; proteose peptone, 0.5% ; $K_2HPO_4$, 0.1% ; $MgSO_4{\cdot}7H_2O$, 0.02% and sodium carbonate, 1.0%. The optimum temperature for alkaline protease production was $35^{\circ}C$, and the initial pH of medium was pH 10.5. The alkaline protease activity was about 2,300 U per ml of culture broth by Casein-Folin Method. A 9.2 fold purification of alkaline protease was obtained from culture broth. The recovery was 14% and purified enzyme was identified as single band, and its molecular weight was about 19,000. The optimum temperature for enzyme reaction was $70^{\circ}C$, and optimum pH was 12. The activity of purified enzyme was inhibited by metal ion ($Fe^{++}$), and Phenylmethylsulfonyl Fluoride, a serine protease inhibitor.

• Journal of Integrative Natural Science
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• v.10 no.3
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• pp.154-161
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• 2017

An antimicrobial peptides-producing Bacillus strain CBS73 was isolated from fermented food (kimchi) that produces low-molecular-weight proteins with broad-spectrum antimicrobial activity. Our goal was to explore the therapeutic potential of antimicrobial substances produced by Bacillus species. Peptide CBS73 was purified from Bacillus subtilis subsp. subtilis with identity of 99.79%. It was found to be stable at pH 4.0-10.0 and temp $20-60^{\circ}C$. A protein band around 5.2 kDa was detected in tricine-SDS-PAGE and band was confirmed by MALDI-TOF test. Peptide CBS73 showed antimicrobial activity against MDR bacteria. The minimal inhibitory concentration (MIC) of peptide CBS73 for vancomycin-resistant S. aureus (VRSA), vancomycin resistant Enterococci (VRE) and Salmonella typhimurium ranged from $10-40{\mu}g/mL$. The antioxidant activity of peptide CBS73 was measured by DPPH scavenging, reducing power activity and total phenolic content. Cell viability and NO production result showed less cytotoxic effect upto $12{\mu}g/mL$. Peptide CBS73 could be a promising antimicrobial agent for clinical application.

• Biotechnology and Bioprocess Engineering:BBE
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• v.8 no.1
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• pp.37-40
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• 2003

The antibacterial action of violet pigment, a mixture of violacein and deoxyviolacein, isolated from phychrotrophic bacterium RT102 strain was examined, and the operational conditions for the effective production of violet pigment were studied. The antibacterial activity of the violet pigment was confirmed for several bacteria such as Bacillus licheniformis, Bacillus subtilis, Bacillus megaterium, Staphylococcus aureus, and Pseudomonas aeruginosa, and the high concentration of violet pigment, above about 15mg/L, caused not only growth inhibition but also death of cells. The growth properties of RT102 strain were clarified under various incubation conditions such as pH, temperature, and dissolved oxygen concentration. The maximum violet pigment concentration, i.e. 3.7 g/L, and the maximum productivity of violet pigment, i.e. 0.12 g .L$\^$-1/H$\^$-1/, were obtained in a batch culture of pH 6, 20$^{\circ}C$, and 1 mg/L of dissolved oxygen concentration.

• 한국생물공학회:학술대회논문집
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• 2005.10a
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• pp.307-311
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• 2005

Marine microorganisms to produce functional biopolymers were isolated from the seashore of the Kyungsang province. Microorganisms to hydrolyze carboxy-methyl cellulose(CMC) were cultured in marin broth and the other liquid medium that contained 2.0% (w/v) glucose, 0.25% yeast extract, 0.5% $K_2HPO_4$, 1% NaCl, 0.02% $MgSO_4{\cdot}7H_2O$ and 0.06% $(NH_4)_2SO_4$ to investigate the ability to produce carboxymethyl cellululase (CMCase) under aerobic conditions. Twelve microorganisms among them showed higher activities of CMCase than B. amyloliquefaciens DL-3, which was known as a cellulase-producing strain. The microorganism showing highest activity of CMCase in this study was identified as Bacillus subtilis subsp. subtilis with 16S rDNA partial sequencing and gyrase A partial sequencing and named as B. subtilis subsp. subtilis A-53.

• Journal of Microbiology and Biotechnology
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• v.31 no.2
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• pp.327-337
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• 2021

Fibrinolytic enzymes with a direct mechanism of action and safer properties are currently requested for thrombolytic therapy. This paper reports on a new enzyme capable of degrading blood clots directly without impairing blood coagulation. This enzyme is also non-cytotoxic and constitutes an alternative to other thrombolytic enzymes known to cause undesired side effects. Twenty-four Bacillus isolates were screened for production of fibrinolytic enzymes using a fibrin agar plate. Based on produced activity, isolate S127e was selected and identified as B. subtilis using the 16S rDNA gene sequence. This strain is of biotechnological interest for producing high fibrinolytic yield and consequently has potential in the industrial field. The purified fibrinolytic enzyme has a molecular mass of 27.3 kDa, a predicted pI of 6.6, and a maximal affinity for Ala-Ala-Pro-Phe. This enzyme was almost completely inhibited by chymostatin with optimal activity at 48℃ and pH 7. Specific subtilisin features were found in the gene sequence, indicating that this enzyme belongs to the BPN group of the S8 subtilisin family and was assigned as AprE127. This subtilisin increased thromboplastin time by 3.7% (37.6 to 39 s) and prothrombin time by 3.2% (12.6 to 13 s), both within normal ranges. In a whole blood euglobulin assay, this enzyme did not impair coagulation but reduced lysis time significantly. Moreover, in an in vitro assay, AprE127 completely dissolved a thrombus of about 1 cc within 50 min and, in vivo, reduced a thrombus prompted in a rat tail by 11.4% in 24 h compared to non-treated animals.

• Preventive Nutrition and Food Science
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• v.14 no.3
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• pp.252-259
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• 2009

We previously isolated Bacillus strains with high fibrinolytic activities (FAs) from cheonggukjang prepared by traditional ways. To test their potential as starters for cheonggukjang, soybean was fermented for 72 hr at $37^{\circ}C$ with each isolate and a control lab strain: B. subtilis CH3-25 (BS3-25), B. amyloliquefaciens CH51 (BA51), B. amyloliquefaciens CH86-1 (BA86-1), and B. subtilis 168 (BS168, control, lab strain). Viable cell numbers of all cheonggukjang samples rapidly increased and reached about $10^9$ CFU/g after 6 hr. During 72 hr, the initial pH of 6.3 rapidly increased to 8.1$\sim$8.2 for cheonggukjang fermented with BS3-25 or BA86-1, and 7.3 for those with BA51 or BS168. FAs and protease activities (acid, neutral, and alkaline) rapidly increased in cheonggukjang fermented with BS3-25, BA51, or BA86-1 during the first 12 hr. On the other hand, those of cheonggukjang fermented with BS168 slightly increased during the first 36 hr. There were significant changes in acid and neutral protease activities in cheonggukjang fermented with BA51 or BA86-1 during the 24 hr. Rapid increases of $\beta$-glucosidase activity corresponded well with rapid increases of $\alpha$-amylase and $\alpha$-galactosidase activities in addition to increases in antioxidant activities and the TPCs (total phenolic contents). The highest increase in the TPCs was observed in cheonggukjang fermented with BA86-1 while the least was that fermented with BS168.