• 한국식품위생안전성학회지
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• 제16권3호
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• pp.188-193
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• 2001

청국장은 전통발효식품으로 우리 식단의 중요한 단백질 공급원으로 자리해 왔으며, 발효과정 중에 나타나는 생리활성기능은 미생물의 작용으로 생성되는 점질물에 의한 것으로 추정된다. 청국장 점질물의 생리활성기능을 확인하기 위하여, 점질물을 첨가하여 유해미생물의 성장곡선을 관찰하였다. 발효균주 Bacillus circulans K-1, Bacillus spp. N-l, Bacillus subtilis CH-1균주를 이용하여 각각의 청국장을 제조한 후, 청국장으로부터 생성된 점질물인 K-1, N-1, CH-1 등을 분리하여 실험에 사용하였다. 청국장 점질물 K-1, N-1, CH-1을 이용하여 항균활성을 측정한 결과, 그람양성세균(Staphylococcus aureus, Bacillus cereus, Bacillus subtilis, Microscopics luteus), 그람음성세균(Escherichia coli O157:H7, Salmonella typhimurium, Pseudomonas fluorescens), 효모(Pichia membyanaefacien, Sacchamyces cerevisiae, Candida albicans)등에 항균작용을 나타내었다. 청국장 점질물은 Bacillus cereus에 대해 80%의 높은 성장억제력을 나타내었으며, Escheyichia coli O157:H7에 대해서는 20%의 약한 성장억제력을 나타내었다. 청국장 점질물 중 CH-1은 세균, 효모에 대해 40%, K-1과 N-1은 20%정도의 성장억제력을 나타내었다.

• 한국양식학회지
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• 제19권1호
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• pp.40-46
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• 2006

Bacillus subtilis strain with highly active chitosanase was isolated from the intestine of Sebastiscus marmoratus (scorpion fish). It was named as Bacillus subtilis CH1 by morphological, biochemical and 165 rRNA gene analysis. The optimal conditions for chitosanase production were investigated. The optimum carbon and nitrogen sources for Bacillus stibtilis CH1 were 2% starch and 1% yeast extract respectively. Unlike other chitosanases, the expression of this chitosanase was not induced or slightly induced with chitosan. The chitosanase secreted into the medium were concentrated with ammonium sulfate precipitation and purified by gel permeation chromatography. The molecular weight of purified chitosanase was 30 kDa. The optimum pH and temperature of purified chitosanase were 5.5 and $60^{\circ}C$ respectively. The purified chitosanase was continuously thermostable at $40^{\circ}C$ and showed stable activity between pH 6.0 and 8.0. Chitosanase activity of Bacillus subtilis CH1 under optimum condition was 4.1 units/ml.

• Current Research on Agriculture and Life Sciences
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• 제31권1호
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• pp.30-37
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• 2013

Bacillus subtilis and Bacillus amyloliquefaciens are closely related species that share a similar genomic background, and are both known to secrete large amounts of proteins directly into a medium. The extracellular proteomes of two strains of Bacillus subtilis and two strains of Bacillus amyloliquefaciens were compared by 2-D gel electrophoresis during the late exponential growth phase. The relative abundance of some minor protein spots varied among the four strains of Bacillus. Over 123 spots of extracellular proteins were visualized on the gel for B. subtilis CH 97, 68 spots for B. subtilis 3-5, 230 spots for B. amyloliquefaciens CH 51, and 60 spotsfor B. amyloliquefaciens 86-1. 2D gel electrophoresis images of the four Bacillus strains showed significantly different protein profiles. Consistent with the 2D gel electrophoretic analysis, most of the B. subtilis proteins differed from the proteases secreted by the B. amyloliquefaciensstrains. Among the proteins identified from B. subtilis, approximately 50% were cytoplasmic and 30% were canonically extracellular proteins. The secreted protein profiles for B. subtilis CH 97 and B. subtilis 3-5 were quite different, as were the profiles for B. amyloliquefaciens CH 51 and 86-1. The four proteomes also differed in the major protein composition. The B. subtilis CH 97 and B. amyloliquefaciens CH 51 proteomes both contained large amounts of secreted hydrolytic enzymes. Among the four strains, B. subtilis 3-5 secreted the least number of proteins. Therefore, even closely related bacteria in terms of genomic sequences can still have significant differences in their physiology and proteome layout.

• 한국식품영양과학회지
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• 제31권3호
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• pp.389-393
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• 2002

청국장에서 분리한 Bacillus subtilis CH-1, Bacillus circul문 K-1과 Bacillus subtilis(natto) N-1 모두 biosurfactant를 생성하며 이 중 Bacillus subtilis CH-1가 가장 큰 생성력을 나타냈다. Biosurfactant를 대량 생산하기 위하여 AM, LM, NB과 TSB 배지중 AM을 기본배지로 선정하여 최적 탄소원과 질소원으로 glucose 2%, soy peptone 0.3%와 무기염을 포함하는 합성배지를 완성하였다. Biosurfactant의 생성은 96시간에 최대를 나타냈으며 이때 배지의 표면장력은 초기값의 약 43% 값을 나타냈다. 한편 배양온도 및 pH는 biosurfactant생산에 크게 영향을 주지 않았으며 pH5.0~8.0범위에서 대체적으로 안정한 생성을 유지하였다. 최적조건에서 배양시 crude biosurfactant 수율은 6 g/L를 얻을수 있다.

• 한국균학회지
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• 제24권4호통권79호
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• pp.293-304
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• 1996

인삼포 토양으로부터 강력한 항진균력을 보이는 KS1 균주를 분리하고 Bacillus subtilis로 동정하였다. 온상시험에서 B. subtilis KS1의 배양액은 오이잿빛곰팡이병, 밀붉은녹병 등의 진균성 농작물병에 대하여 탁월한 방제효과를 나타내었다. 또한, B. subtilis KS1 배양액의 butanol 분획은 Botrytis maydis, Chytridium lagenarium, Candida albicans 등 식물 또는 사람의 병원성 진균을 비롯한 몇몇 진균류에 대하여 억제효과를 나타내었다. 이 균주가 생성하는 항진균물질을 pep-RPC reverse phase column과 ${\mu}$ Bondapak $C_{18}$ reverse phase column을 이용하여 분리 정제하였다. 정제된 항진균물질의 열안정성 및 pH 안정성을 조사한 결과, $-20-121^{\circ}C$와 pH 4.0-10.0의 범위에서 안정하였다. 이 물질의 조성 및 구조적 특징을 HPLC와 $^1H-,\;^1H-^1H-$, COSY, NOESY, COSY-NOESY 및 HOHAHA NMR spectroscopy를 통하여 각각 분석한 결과, B. subtilis가 생산하는 고리형 peptide 중 iturin A에 속하는 물질로 확인되었다. 그러나, 지금까지 알려진 iturin A계 물질들의 ${\beta}$-아미노산에 붙어 있는 탄화수소 사슬이 곁가지가 없거나 terminus 또는 subterminus에 1개의 $CH_3$ 곁가지를 갖는 것과는 달리, SW1의 ${\beta}$-아미노산은 2개의 $-CH_3$ 곁가지가 붙어 있는 탄화수소 사슬을 가짐이 확인되었다.

• Journal of Microbiology and Biotechnology
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• 제20권2호
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• pp.370-374
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• 2010

A gene encoding the major secreted fibrinolytic protein of Bacillus amyloliquefaciens CH86-1 was cloned from genomic DNAs. DNA sequencing showed that the gene, aprE86-1, could direct the synthesis of a mature protein 275 amino acids in length after processing. When aprE86-1 was introduced into B. subtilis, a mature 27-kDa protein was produced as expected. The fibrinolytic activity of the B. subtilis transformant (TF) was higher than that of B. amyloliquefaciens CH86-1, showing the possibility of increasing the fibrinolytic activity of Bacillus strains through genetic engineering.

• 한국양식학회:학술대회논문집
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• 한국양식학회 2006년도 수산관련학회 공동학술대회 발표요지집
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• pp.279-280
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• 2006

• Preventive Nutrition and Food Science
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• 제14권3호
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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.

• 한국유기농업학회지
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• 제7권2호
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• pp.99-106
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• 1999

사과나무에서 발생하는 탄저병에 대한 길항미생물을 찾기 위하여 자연계로부터 유용미생물을 분리하고 사과 탄저 병원균에 대한 길항력 검정과 균주를 동정한 결과는 다음과 같다. 자연계로부터 얻은 1,000여종의 미생물중에서 탄저병원균에 대하여 길항력이 우수한 미생물을 1차적으로 11종 선발하였으며, 이중에서 가장 길항력이 뛰어난 미생물 CH1141을 최종적으로 선발하였다. 길항미생물 CH1141은 분리한 탄저병원균에 대하여 65%의 높은 생장억제력을 보였다. 길항력이 우수한 CH1141의 형태적 성질, 배양적 특성 및 생리 생화학적 성질 등을 조사하여 비교 검토한 결과 Bacillus subtilis와 유사한 균으로 동정되었다.

• Journal of Microbiology and Biotechnology
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• 제31권6호
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• pp.833-839
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• 2021

Bacillus subtilis CH3-5 isolated from cheonggukjang secretes a 28 kDa protease with a strong fibrinolytic activity. Its gene, aprE3-5, was cloned and expressed in a heterologous host (Jeong et al., 2007). In this study, the promoter of aprE3-5 was replaced with other stronger promoters (P_cry3A, P₁₀, P_SG1, P_srfA) of Bacillus spp. using PCR. The constructed chimeric genes were cloned into pHY300PLK vector, and then introduced into B. subtilis WB600. The P10 promoter conferred the highest fibrinolytic activity, i.e., 1.7-fold higher than that conferred by the original promoter. Overproduction of the 28 kDa protease was confirmed using SDS-PAGE and fibrin zymography. RT-qPCR analysis showed that aprE3-5 expression was 2.0-fold higher with the P10 promoter than with the original promoter. Change of the initiation codon from GTG to ATG further increased the fibrinolytic activity. The highest aprE3-5 expression was observed when two copies of the P₁₀ promoter were placed in tandem upstream of the ATG initiation codon. The construct with P10 promoter and ATG and the construct with two copies of P10 promoter in tandem and ATG exhibited 117% and 148% higher fibrinolytic activity, respectively, than that exhibited by the construct containing P10 promoter and GTG. These results confirmed that significant overproduction of a fibrinolytic enzyme can be achieved by suitable promoter modification, and this approach may have applications in the industrial production of AprE3-5 and related fibrinolytic enzymes.