• Journal of Microbiology and Biotechnology
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• 제13권3호
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• pp.451-456
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• 2003

An expression vector system was developed for the secretory production of recombinant Bacillus stearothermophilus L1 lipase in Saccharomyces cerevisiae. The mature L1 lipase gene was fused to ${\alpha}-amylase$ signal sequence from Aspergillus oryzae for the effective secretion into the culture broth and the expression was controlled under GAL10 (the gene coding UDP-galactose epimerase of S. cerevisiae) promoter. S. cerevisiae harboring the resulting plasmid successfully secreted L1 lipase into the culture broth. To examine an optimum condition for L1 lipase expression in the fed-batch culture, L1 lipase expression was induced at three different growth phases (early, mid, and late-exponential growth phases). Maximum product on of L1 lipase (1,254,000 U/l, corresponding to 0.65/1) was found when the culture was induced at an early growth phase. Secreted recombinant L1 lipase was purified only through CM-Sepharose chromatography, and the purified enzyme showed 1,963 U/mg of specific activity and thermoalkalophilic properties similar to those reported for the enzyme expressed in Escherichia coli.

• Journal of Microbiology and Biotechnology
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• 제14권3호
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• pp.532-539
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• 2004

For efficient lincomycin production from Streptomyces lincolnensis L1245, various vegetable oils, natural nitrogen sources, and surfactants were investigated at the pilot-scale level in the flask. Olive oil as the sole carbon source was the most suitable one for producing lincomycin. When 20 g/lof olive oil was used, the lincomycin concentration and lipase activity reached 1.01 g/land 182 U/ml, respectively, after 5 days of culture. Among the various unsaturated fatty acids, when linolenic acid was used, the cell growth and lincomycin production were markedly decreased. On the other hand, when 0.2 g/l of oleic acid was added to the culture broth, the maximum lincomycin concentration was 1.0 g/l, which was about 1.7-fold higher than that obtained without the addition of oleic acid. Among the various natural nitrogen sources, pharmamedia or soybean meal was the most suitable nitrogen source. In particular, in the case of a mixture of 10 g/l of pharmamedia and soybean meal, 1.5 g/l of lincomycin concentration and 220 U/ml of lipase activity were obtained. When Span 180 was used as the surfactant, lincomycin production, lipase activity, and oil consumption increased. The correlation between the consumption rates of oil and lincomycin production in a culture using olive oil as the sole carbon source was also investigated. The lincomycin production depended on the consumption rate of olive oil. Using these results, fed-batch cultures for comparing the use of olive oil and starch as a conventional carbon source were carried out in a 5-1 fermentor. When olive oil was used as the sole carbon source, 34 g/l of olive oil was consumed after 7 days of culture. The maximum lincomycin concentration was 3.0 g/l, which was about 2.0-fold higher than that of starch medium after 7 days of culture. The product yield was 0.09 gig of consumed carbon source, which was about 3.0-fold higher than that of starch medium after 7 days of culture.

• Journal of Microbiology and Biotechnology
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• 제25권11호
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• pp.1819-1826
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• 2015

Poly(ε-ʟ-lysine) (ε-PL) is a novel bioactive polymer secreted by filamentous bacteria. Owing to lack of a genetic system for most ε-PL-producing strains, very little research on enhancing ε-PL biosynthesis by genetic manipulation has been reported. In this study, an effective genetic system was established via intergeneric conjugal transfer for Streptomyces albulus PD-1, a famous ε-PL-producing strain. Using the established genetic system, the Vitreoscilla hemoglobin (VHb) gene was integrated into the chromosome of S. albulus PD-1 to alleviate oxygen limitation and to enhance the biosynthesis of ε-PL in submerged fermentation. Ultimately, the production of ε-PL increased from 22.7 g/l to 34.2 g/l after fed-batch culture in a 5 L bioreactor. Determination of the oxygen uptake rate, transcriptional level of ε-PL synthetase gene, and ATP level unveiled that the expression of VHb in S. albulus PD-1 enhanced ε-PL biosynthesis by improving respiration and ATP supply. To the best of our knowledge, this is the first report on enhancing ε-PL production by chromosomal integration of the VHb gene in an ε-PL-producing strain, and it will open a new avenue for ε-PL production.

• Journal of Microbiology and Biotechnology
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• 제20권1호
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• pp.101-109
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• 2010

Efficient L-lactic acid production from Jerusalem artichoke tubers, by Lactobacillus casei G-02, using simultaneous saccharification and fermentation (SSF) in a fed-batch culture, is demonstrated. A kinetic analysis of the SSF revealed that the inulinase activity was subjected to product inhibition, whereas the fermentation activity of G-02 was subjected to substrate inhibition. It was also found that the intracellular NADH oxidase (NOX) activity was enhanced by the citrate metabolism, which dramatically increased the carbon flux of the Embden-Meyerhof-Parnas (EMP) pathway, along with the production of ATP. As a result, when the SSF was carried out at $40^{\circ}C$ after an initial hydrolysis of 1 h and included a sodium citrate supplement of 10 g/l, an L-lactic acid concentration of 141.5 g/l was obtained after 30 h, with a volumetric productivity of 4.7 g/l/h. The conversion efficiency and product yield were 93.6% of the theoretical lactic acid yield and 52.4 g lactic acid/l00 g Jerusalem artichoke flour, respectively. Such a high concentration of lactic acid with a high productivity from Jerusalem artichokes has not been reported previously, making G-02 a potential candidate for the economic production of L-lactic acid from Jerusalem artichokes on a commercial scale.

• Journal of Microbiology and Biotechnology
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• 제24권6호
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• pp.781-787
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• 2014

2-Ketogluconic acid production by Klebsiella pneumoniae is a pH-dependent process, strictly proceeding under acidic conditions. Unfortunately, cell growth is inhibited by acidic conditions, resulting in low productivity of 2-ketogluconic acid. To overcome this deficiency, a two-stage fermentation strategy was exploited in the current study. During the first stage, the culture was maintained at neutral pH, favoring cell growth. During the second stage, the culture pH was switched to acidic conditions favoring 2-ketogluconic acid accumulation. Culture parameters, including switching time, dissolved oxygen levels, pH, and temperature were optimized for the fed-batch fermentation. Characteristics of glucose dehydrogenase and gluconate dehydrogenase were revealed in vitro, and the optimal pHs of the two enzymes coincided with the optimum culture pH. Under optimum conditions, a total of 186 g/l 2-ketogluconic acid was produced at 26 h, and the conversion ratio was 0.98 mol/mol. This fermentation strategy has successfully overcome the mismatch between optimum parameters required for cell growth and 2-ketogluconic acid accumulation, and this result has the highest productivity and conversion ratio of 2-ketogluconic and produced by microorganism.

• Journal of Microbiology and Biotechnology
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• 제30권1호
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• pp.136-145
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• 2020

Chlorella sp. HS2, which previously showed excellent performance in phototrophic cultivation and has tolerance for wide ranges of salinity, pH, and temperature, was cultivated heterotrophically. However, this conventional medium has been newly optimized based on a composition analysis using elemental analysis and ICP-OES. In addition, in order to maintain a favorable dissolved oxygen level, stepwise elevation of revolutions per minute was adopted. These optimizations led to 40 and 13% increases in the biomass and lipid productivity, respectively (7.0 and 2.25 g l^-1d^-1 each). To increase the lipid content even further, 12 h heat shock at 50℃ was applied and this enhanced the biomass and lipid productivity up to 4 and 17% respectively (7.3 and 2.64 g l^-1d^-1, each) relative to the optimized conditions above, and the values were 17 and 14% higher than ordinary lipid-accumulating N-limitation (6.2 and 2.31 g l^-1d^-1). On this basis, heat shock was successfully adopted in novel Chlorella sp. HS2 cultivation as a lipid inducer for the first time. Considering its fast and cost-effective characteristics, heat shock will enhance the overall microalgal biofuel production process.

• Journal of Microbiology and Biotechnology
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• 제10권1호
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• pp.75-80
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• 2000

An improved method of refolding recombinant human proinsulin from E. coli was presented. It was based on a two-stage stirred tank reactor in which denatured proinsulin-s-sulfonate was mixed instantaneously with a reaction buffer in the first stage reactor, and then fed to the second stage reactor. The mixture was stirred further for a total of 30h in the second stage reactor. In this system, unfavorable effects present due to the increase in reaction volume and protein concentration for protein refolding, which becomes significant in a large-scale operation, were avoided. Refolding yields of over 80% was obtained for achieving reaction volume of upto 50 l at protein concentration of 1 mg/ml. The optimum urea concentration was 1M. Refolding yield at the 1-1 reaction volume and protein concentration of 0.5mg/ml was increased about 2.5-fold, compared to that in a batch reactor. By increasing protein concentration in a two-stage refolding reaction, the cost for insulin production could be reduced, therefore, making this process economical.

• Journal of Microbiology and Biotechnology
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• 제27권6호
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• pp.1112-1119
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• 2017

The aim of this study was to develop a potential process for bioethanol production from Hydrodictyon reticulatum (HR), a filamentous freshwater alga, using Saccharomyces cerevisiae (KCTC7017). From the sugar solutions prepared by the four different hydrolysis methods, bioethanol production ranged from 11.0 g/100 g dried material (acid hydrolysis) to 22.3 g/100 g dried material (enzymatic hydrolysis, EH). Bioethanol was fermented from a highly concentrated sugar solution obtained by a decompression-mediated (vacuum) enrichment method (VE). As the results, ethanol was more efficiently produced from HR when sugar solutions were concentrated by VE following EH (EH/VE). Using multiple feeding of the sugar solution prepared by EH/VE from HR, ethanol reached up to a concentration of 54.3 g/l, corresponding to 24.9 g/100 g dried material, which attained the economic level of product concentration (approximately 5%). The results indicate that by using HR, it is feasible to establish a bioethanol production process, which is effective for using microalgae as the raw material for ethanol production.

• Journal of Microbiology and Biotechnology
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• 제17권7호
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• pp.1071-1078
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• 2007

Using a 50-1 airlift bioreactor, for the effective production of tylosin from Streptomyces fradiae TM-224 using raw cornmeal as the energy source, various environmental factors were studied in flask cultures. The maximum tylosin concentration was obtained at $32^{\circ}C$ and pH between 7.0 and 7.5. When seed was inoculated after 24 h of culture, the maximum tylosin concentration, 5.7 g/l, was obtained after 4 days of culture. Various concentrations of raw cornmeal were tested to investigate the optimum initial concentration for the tylosin production. An initial raw cornmeal concentration of 80 g/l gave the highest tylosin concentration, 5.8 g/l, after 5 days of culture. Of the various nitrogen sources, soybean meal and fish meal were found to be the most effective for the production of tylosin. In particular, with the optimal mixing ratio, 12 g/l of soybean meal to 14 g/l of fish meal, 7.2 g/l of tylosin was obtained after 5 days of culture. To compare raw cornmeal and glucose for the production of tylosin in the 50-1 airlift bioreactor for 10 days, fed-batch cultures were carried out under the optimum culture conditions. When raw com meal was used as the energy source, the tylosin production increased with increasing culture time. The maximum tylosin concentration after 10 days of culture was 13.5 g/l, with a product yield from raw cornmeal of 0.123 g/g of consumed carbon source, which was about 7.2 times higher than that obtained when glucose was used as the carbon source.

• Journal of Microbiology and Biotechnology
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• 제22권9호
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• pp.1258-1263
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• 2012

2,3-Butanediol (2,3-BD) is a major metabolite produced by Klebsiella pneumoniae KCTC2242, which is a important chemical with wide applications. Three genes important for 2,3-BD biosynthesis acetolactate decarboxylase (budA), acetolactate synthase (budB), and alcohol dehydrogenase (budC) were identified in K. pneumoniae genomic DNA. With the goal of enhancing 2,3-BD production, these genes were cloned into pUC18K expression vectors containing the lacZ promoter and the kanamycin resistance gene to generate plasmids pSB1-7. The plasmids were then introduced into K. pneumoniae using electroporation. All strains were incubated in flask experiments and 2,3-BD production was increased by 60% in recombinant bacteria harboring pSB04 (budA and budB genes), compared with the parental strain K. pneumoniae KCTC2242. The maximum 2,3-BD production level achieved through fed-batch fermentation with K. pneumoniae SGJSB04 was 101.53 g/l over 40 h with a productivity of 2.54 g/l.h. These results suggest that overexpression of 2,3-BD synthesis-related genes can enhance 2,3-BD production in K. pneumoniae by fermentation.