• Applied Biological Chemistry
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• 제48권4호
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• pp.345-351
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• 2005

난배양 미생물로부터 천연물질을 찾기 위하여 토양으로부터 직접분리 된 DNA와 cosmid vector를 이용하여 metagenomic library를 제작하고 탐색 하였다. 대장균에서 발현되는 유전자은행 초기 탐색 결과 LB배지에서 잘 자라면서 브라운 색깔을 내는 여러 개의 clone을 선발 하였다. 선발된 여러 후보 clone중 pYS85C는 돌연변이를 유도하였으며 색깔을 생산하지않는 clone 들에 대하여 염기서열을 결정 하였다. 돌연변이clone들로부터 결정된 pYS85C 염기서열 결과 아미노산이 393개이며 44.5 kDa으로 색소형성에 관여하는 4-hydroxyphenylpyruvic acid dioxygenase(HPPD) 유전자로 판명 되었다. 또한, BLAST비교 분석에서 이효소는 기존에 밝혀진 HPPD효소와 60% 정도의 identity를 보였고 C-말단에서는 많은 conserved domain이 있었다. 이러한 결과로 볼 때 천연물질을 합성 할 수 있는 유전자는 토양DNA로부터 직접 분리되어 발현될 수 있으며 이러한 기술은 새로운 물질을 찾는데 중요한 tool이 될 수 있다.

• Journal of Microbiology and Biotechnology
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• 제17권6호
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• pp.905-912
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• 2007

A novel ${\beta}-glucosidase$ gene, bglA, was isolated from uncultured soil bacteria and characterized. Using genomic libraries constructed from soil DNA, a gene encoding a protein that hydrolyzes a fluorogenic analog of cellulose, 4-methylumbelliferyl ${\beta}-D-cellobioside$ (MUC), was isolated using a microtiter plate assay. The gene, bglA, was sequenced using a shotgun approach, and expressed in E. coli. The deduced 55-kDa amino acid sequence for bglA showed a 56% identity with the family 1 glycosyl hydrolase Chloroflexus aurantiacus. BglA included two conserved family 1 glycosyl hydrolase regions. When using $p-nitrophenyl-{\beta}-D-glucoside$ (pNPG) as the substrate, the maximum activity of the purified ${\beta}-glucosidase$ exhibited at pH 6.5 and $55^{\circ}C$, and was enhanced in the presence of $Mn^{2+}$. The $K_m\;and\;V_{max}$ values for the purified enzyme with pNPG were 0.16 mM and $19.10{\mu}mol/min$, respectively. The purified BglA enzyme hydrolyzed both pNPG and $p-nitrophenyl-{\beta}-D-fucoside$. The enzyme also exhibited substantial glycosyl hydrolase activities with natural glycosyl substrates, such as sophorose, cellobiose, cellotriose, cellotetraose, and cellopentaose, yet low hydrolytic activities with gentiobiose, salicin, and arbutin. Moreover, BglA was able to convert the major ginsenoside $Rb_1$ into the pharmaceutically active minor ginsenoside Rd within 24 h.

• 한국미생물·생명공학회지
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• 제40권1호
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• pp.30-38
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• 2012

두 종의 Bacillus와 한 종의 Pseudomonas로 구성된 컨소시엄 미생물제제를 고추 경작지에 첨가하였을 때 경작지 토양의 미생물 다양성 변화를 조사하기 위해 건전한 토양의 지표가 되는 질소순환관여 미생물과 근권토양 내 섬유소 분해기능, 인산가용화기능과 urea 가수분해기능을 가지는 기능성미생물의 활성과 밀도를 측정하였다. 고추 경작지 토양에 컨소시엄 PGPR 미생물인 B. subtilis AH18, B. licheniformis K11과 P. fluorescens 2112의 컨소시엄 미생물제제를 처리하여 토양미생물의 다양성에 변화를 측정하여 컨소시엄 미생물제제의 처리에 의한 근권토양 미생물의 다양성 변화와 고추의 생육에 미치는 영향을 분석하였다. 컨소시엄 미생물제제 처리구의 경우 Actinomyces spp., Trichoderma spp., 광합성세균과 Azotobacter spp.가 다른 처리구에 비하여 1.7-5배 이상 높았으며, 질소 순환에 관여하는 미생물은 1.4-4배 증가되었다. 또한 기능성 미생물의 다양성도 화학제제나 물처리구와 비교하였을 때 1.3-3배 증가하였다. 건고추 수확량은 컨소시엄 미생물제제를 처리하였을 경우 대조구나 화학제제를 처리하였을 때보다 15%이상 수확량이 증대되었다. 따라서 PGPR 컨소시엄 미생물제제의 처리로 인해 고추 경작지 토양의 근권미생물 다양성과 작물의 생산성에 긍정적인 영향을 주는 것으로 생각된다. 추후 메타지노믹스를 활용하여 미생물 다양성 변화를 추가적으로 확인할 예정이다.

• 한국미생물·생명공학회지
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• 제51권1호
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• pp.124-127
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• 2023

Among certain animals, gut microbiomes demonstrate species-specific patterns of beta diversity. This host-specificity is a potent driver of exogenous microbial exclusion. To overcome persistent translational limitations, translational microbiome research and therapeutic development must account for host-specific patterns of microbial engraftment. This commentary seeks to highlight the important implications of host-specificity for microbial ecology, Fecal Microbiota Transplantation (FMT), next-generation probiotics, and translational microbiota research.

• 한국미생물·생명공학회지
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• 제24권1호
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• pp.1-8
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• 1996

Characterization and numerical identification were carried out for an actinomycetes SL20209. Morphological, cultural and physiological perperties of SL20209 which porduced valistatin and des-$asp^4$-amastatin as inhibitors of aminopeptidase M were evaluated. The isolate was identified to be the genus of Streptomyces. Fourty-three taxonomic units were analysed by using a TAXON program. The isolate was classified into the major cluster 29 of Streptomyces and best-matched to Streptomyces griseoplanus.

• Journal of Microbiology and Biotechnology
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• 제21권1호
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• pp.81-88
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• 2011

The anti-phytoplasma activities of surfactin (derived from Iranian native Bacillus subtilis isolates) and tetracycline towards Candidatus "Phytoplasma aurantifolia", the agent of lime Witches' broom disease, were investigated. HPLC was used to quantify the surfactin production in four previously characterized native surfactin-producing strains, and the one producing the highest amount of surfactin (about 1,500 mg/l) was selected and cultivated following optimized production and extraction protocols. Different combinations of purified surfactin and commercial tetracycline were injected into artificially phytoplasmainfected Mexican lime seedlings using a syringe injection system. An absolute quantitative real-time PCR system was developed to monitor the phytoplasma population shifts in the lime phloem during 3 months following the injections. The results revealed that the injections of surfactin or tetracycline had a significant inhibitory effect on Candidatus "P. aurantifolia". However, the combined treatment with both surfactin and tetracycline (1:1) resulted in the highest inhibition due to a synergic effect, which suppressed the phytoplasma population from about $2{\times}10^5$ to less than 10 phytoplasma units/g plant tissue.

• Journal of Microbiology and Biotechnology
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• 제6권3호
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• pp.213-218
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• 1996

Hot-water extract, Fr. 1, of Phellinus linteus mycelium was fractionated into Fr. 2, 3, 4, and 5 by the difference of solubility in ethanol. The polysaccharide fractions were studied for their immunostimulating activity on in vitro T-independent polyc1onal antibody response to trinitrophenyl-haptened SRBC (sheep red blood cell). The Fr. 4 with the highest immunostimulating activity was subjected to DEAE-cellulose ion exchange chromatography and gave five fractions, 4-I, II, III, IV, and V. The in vitro immunostimulating assay of the five fractions showed that 4-I and 4-III had a similar activity to that of LPS but the other fractions had low activity. By analyses of chemical composition and HPLC, all fractions obtained were found to be heteropolysaccharide-protein complex. The molecular weights ranged from 9, 000 to 15, 000. Sugar analyses showed that glucose, galactose, mannose, arabinose, and xylose were main component. Uronic acid and amino sugar were also detected in the fractions. It should be noted that the molecular weight (15, 000) of 4-III was very small and the structure of 4-III may be different from the known immunostimulating branched $\beta$-(1longrightarrow3)-glucan.

• Journal of Microbiology and Biotechnology
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• 제13권6호
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• pp.892-896
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• 2003

High-level expression of the lipase B26 gene from Bacillus pumilus was achieved using Bacillus subtilis Chungkookjang isolated from the Korean traditional fermented bean paste, Chungkookjang. For the secretory production of recombinant lipase B26 in a Bacillus host system, pLipB26 was constructed by ligating the lipase B26 gene into the recently designed Escherichia coli-Bacillus shuttle vector, pLipSM, and that was then transformed into B. subtilis Chungkookjang. Among the various vector, medium, and host combinations, B. subtilis Chungkookjang harboring the pLipB26 exhibited the highest lipase activity in PY medium, and B. subtilis Chungkookjang secreted two times more enzymes than B. subtilis DB 104 under the same condition. When B. subtilis Chungkookjang harboring the pLipB26 was cultured in a 5-1 jar-fermentor containing 21 of a PY medium, the maximum lipase activity (140 U/ml) and production yield (0.68 g/l) were obtained during the late exponential phase from a cell-free culture broth. Although B. subtilis Chungkookjang also secreted extracellular proteases at the late exponential phase, these results suggested the potential of B. subtilis Chungkookjang as a host for the secretory production of foreign proteins.

• Journal of Microbiology and Biotechnology
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• 제23권5호
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• pp.674-680
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• 2013

Yeast Dekkera/Brettanomyces bruxellensis is probably the most common contaminant in wineries and ethanol production processes. The considerable economic losses caused by this yeast, but also its ability to produce and tolerate high ethanol concentrations, make it an attractive subject for research with potential for industrial applications. Unfortunately, efforts to understand the biology of D. bruxellensis and facilitate its broader use in industry are hampered by the lack of adequate procedures for delivery of exogenous DNA into this organism. Here we describe the development of transformation protocols (spheroplast transformation, LiAc/PEG method, and electroporation) and report the first genetic transformation of yeast D. bruxellensis. A linear heterologous DNA fragment carrying the kanMX4 sequence was used for transformation, which allowed transformants to be selected on plates containing geneticin. We found the spheroplast transformation method using 1M sorbitol as osmotic stabilizer to be inappropriate because sorbitol strikingly decreases the plating efficiency of both D. bruxellensis spheroplast and intact cells. However, we managed to modify the LiAc/PEG transformation method and electroporation to accommodate D. bruxellensis transformation, achieving efficiencies of 0.6-16 and 10-20 transformants/${\mu}g$ DNA, respectively. The stability of the transformants ranged from 93.6% to 100%. All putative transformants were analyzed by Southern blot using the kanMX4 sequence as a hybridization probe, which confirmed that the transforming DNA fragment had integrated into the genome. The results of the molecular analysis were consistent with the expected illegitimate integration of a heterologous transforming fragment.

• 한국미생물·생명공학회지
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• 제49권4호
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• pp.501-509
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• 2021

The increasing demand for baked products has given a boost to research on isolation and selection of novel yeast strains with improved leavening activity. Twelve sourdough samples were collected from several localities of the Fez region in Morocco. The pH and total titratable acidity (TTA) values of these samples varied from 3.03-4.63 and 14-17.5 ml of 0.1 N NaOH/10 g of sourdough, respectively, while yeast counts ranged from 5.3 6.77 Log CFU/g. Thirty-two yeast isolates were obtained and evaluated for their leavening ability. Out of all isolates, four yeasts molecularly identified as Saccharomyces cerevisiae (three strains) and Kluyveromyces marxianus (one strain) showed highest specific volumes of 4.69, 4.55, 4.35 and 4.1 cm³/g, respectively. These strains were further assessed for their tolerance to high concentrations of salt, sugar, elevated temperatures, and low pH conditions. K. marxianus showed higher resistance than the S. cerevisiae. Thus, Moroccan sourdoughs harbor technologically relevant yeasts that could be used as potential starters for bread preparation.