• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.666-674
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• 2016

The bacterial strains were screened as potential starters for fermenting low-salt doenjang (a Korean traditional fermented soybean paste) using Korean doenjang based on proteolytic and antipathogenic activities under 6.5-7.5% NaCl conditions. Phylogenetic analysis based on 16S rRNA gene sequences showed that they all belonged to the genus Bacillus. Proteolytic and antipathogenic activities against Escherichia coli, Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, and Aspergillus flavus, as well as fibrinolytic, amylase, and cellulase activities of the 10 strains were quantitatively evaluated. Of these, strains D2-2, JJ-D34, and D12-5 were selected, based on their activities. The functional, phenotypic, and safety-related characteristics of these three strains were additionally investigated and strains D2-2 and D12-5, which lacked antibiotic resistance, were finally selected. Strains D2-2 and D12-5 produced poly-γ-glutamic acid and showed various enzyme activities, including α-glucosidase and β-glucosidase. Growth properties of strains D2-2 and D12-5 included wide temperature and pH ranges, growth in up to 16% NaCl, and weak anaerobic growth, suggesting that they facilitate low-salt doenjang fermentation. Strains D2-2 and D12-5 were not hemolytic, carried no toxin genes, and did not produce biogenic amines. These results suggest that strains D2-2 and D12-5 can serve as appropriate starter cultures for fermenting low-salt doenjang with high quality and safety.

• Korean Journal of Microbiology
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• v.25 no.2
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• pp.157-164
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• 1987

A new endo-$\beta$-1, 4-glucanase was purified from the culture filtrate of thermophilic anaerobic Clostridium thermocellum. The purification procedure included two steps of ion exchange chromatography with DEAD-Sephadex A-50 and gel filtration chromatography with Sephadex G-75. Even though the 56 fold increase in CMCase specific activity was obtained, the actually recovered enzyme activity was relatively lower level of 0.7%. Judging from the two bands in SDS-polyacrylamide gel electrophoresis, the endo-$\beta$-1, 4-glucanase consists of two subunits whose M.W. are 38,000 and 58,000, respectively. The optimum pH and temperature were determined to be 5.0 and $65^{\circ}C$, respectively. The enzyme was stable up to $70^{\circ}C$, but inactivated at $80^{\circ}C$. The kinetic parameters of the separated fraction were also determined. The purified enzyme did not show any significant hydrolytic activity against the highly ordered crystalline cellulose as well as filter paper.

• Tuberculosis and Respiratory Diseases
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• v.78 no.3
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• pp.289-292
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• 2015

Lemierre syndrome (LS) is a septic thrombophlebitis of the internal jugular vein (IJV) following an oropharyngeal infection. LS is commonly caused by normal anaerobic flora and treated with appropriate antibiotics and anticoagulation therapy. Although the incidence of disease is very rare, 15% cases of LS are fatal even in the antibiotic era because of disseminated septic thromboemboli. We reported a case of extensive bilateral LS due to methicillin-resistant Staphylococcus epidermidis in a 63-year-old female with lung adenocarcinoma. Initial examination revealed a retropharyngeal abscess; hence, intravenous ceftriaxone and steroid were initiated empirically. However, pulmonary thromboembolism developed and methicillin-resistant S. epidermidis was identified in the bacterial culture. Despite intensive antibiotic and anticoagulation therapies, extensive septic thrombophlebitis involving the bilateral IJV and superior vena cava developed. Adjunctive catheter-directed thrombolysis and superior vena cava stenting were performed and the patient received antibiotic therapy for an additional 4 weeks, resulting in complete recovery.

• KSBB Journal
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• v.29 no.3
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• pp.131-138
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• 2014

In this study, we isolated two novel chitinase producing bacterial strains from yellow loess samples collected from Jullanamdo province. The chitinase producing bacteria were isolated based on the zone size of clearance in the chitin agar plates. Both of them were gram positive, rod ($2{\sim}3{\times}0.3{\sim}0.4{\mu}m$), spore-forming, and motility positive. They were facultative anaerobic, catalase positive and hydrolyzed starch, gelatin, and casein. From the 16s rRNA gene sequence analysis, the isolates were labeled as Bacillus licheniformis KYLS-CU01 and B. licheniformis KYLS-CU02. The isolates showed higher extracellular chitinase activities than B. licheniformis ATCC 14580 as a control. The optimum temperature and pH for chitinase production were $40^{\circ}C$ and pH 7.0, respectively. Response Surface Methodology (RSM) was used to optimize the culture medium for efficient production of the chitinase. Under this optimal condition, 1.5 times higher chitinase activity of B. licheniformis KYLS-CU02 was obtained. Extracellular chitinases of the two isolates were purified through ammonium sulfate precipitation and anion-exchange DEAE-cellulose column chromatography. The specific activities of purified chitinase from B. licheniformis KYLS-CU01 and B. licheniformis KYLS-CU02 were 7.65 and 5.21 U/mg protein, respectively. The molecular weights of the two purified chitinases were 59 kDa. Further, the purified chitinase of B. licheniformis KYLS-CU01 showed high antifungal activity against Fusarium sp.. In conclusion, these two bacterial isolates can be used as a biopesticide to control pathogenic fungi.

• Biomolecules & Therapeutics
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• v.15 no.4
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• pp.261-265
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• 2007

Hydroxyproline (HYP) is a post-translational product of proline hydroxylation catalyzed by an enzyme prolyl 4-hydroxylase (P4H) which plays a crucial role in the synthesis of all collagens. Considering the role of collagen and its significance in many clinically important diseases such as liver fibrosis, a great deal of attention has been directed toward the development of an assay at cell-based system. The reason is that cell-based assay system is more efficient than enzyme-based in vitro system and takes much less time than in vivo system. Several assay procedures developed for P4H are laborious, time-consuming and not feasible for the massive-screening. Here, we report the cell-based assay method of prolyl 4-hydroxylase in immortalized rat hepatic stellate HSC-T6 cells. To optimize the cell culture condition to assay for HYP content, various concentrations of reagents were treated for different times in HSC-T6 cells. Our data showed that the treatment with ascorbate in a hypoxic condition for 24 h resulted in the maximal increase of HYP by 1.8 fold. Alternatively, cobalt chloride ($5\;{\mu}M$) and ascorbate ($50\;{\mu}M$) in normoxic states exhibited similar effect on the production of HYP as in a hypoxic condition. Therefore, cobalt chloride can be substituted for a hypoxic condition when an anaerobic chamber is not available. Rosiglitazone and HOE077, known as inhibitors of collagen, synthesis decreased P4H enzyme activity by 32.3% and 15%, respectively, which coincided with previous reports from liver tissues. The level of the smooth muscle ${\alpha}$-actin, a marker of activated stellate cells, was significantly increased under hypoxia, suggesting that our experimental condition could work for screening the anti-fibrotic compounds. The assay procedure took only 3 days after treatment with agents, while assays from the primary stellate cells or liver tissues have taken several weeks. Considering the time and expenses, this assay method could be useful to screen the compounds for the inhibitor of prolyl 4-hydroxylase.

• Asian-Australasian Journal of Animal Sciences
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• v.26 no.4
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• pp.523-528
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• 2013

This study investigates the effects of three branched-chain amino acids (BCAA; valine, leucine, and isoleucine) on the in vitro ruminal fermentation of wheat straw using batch cultures of mixed ruminal microorganisms. BCAA were added to the buffered ruminal fluid at a concentration of 0, 2, 4, 7, or 10 mmol/L. After 72 h of anaerobic incubation, pH, volatile fatty acids (VFA), and ammonia nitrogen ($NH_3$-N) in the ruminal fluid were determined. Dry matter (DM) and neutral detergent fiber (NDF) degradability were calculated after determining the DM and NDF in the original material and in the residue after incubation. The addition of valine, leucine, or isoleucine increased the total VFA yields ($p{\leq}0.001$). However, the total VFA yields did not increase with the increase of BCAA supplement level. Total branched-chain VFA yields linearly increased as the supplemental amount of BCAA increased (p<0.001). The molar proportions of acetate and propionate decreased, whereas that of butyrate increased with the addition of valine and isoleucine (p<0.05). Moreover, the proportions of propionate and butyrate decreased (p<0.01) with the addition of leucine. Meanwhile, the molar proportions of isobutyrate were increased and linearly decreased (p<0.001) by valine and leucine, respectively. The addition of leucine or isoleucine resulted in a linear (p<0.001) increase in the molar proportions of isovalerate. The degradability of NDF achieved the maximum when valine or isoleucine was added at 2 mmol/L. The results suggest that low concentrations of BCAA (2 mmol/L) allow more efficient regulation of ruminal fermentation in vitro, as indicated by higher VFA yield and NDF degradability. Therefore, the optimum initial dose of BCAA for in vitro ruminal fermentation is 2 mmol/L.

• Journal of the Korean Applied Science and Technology
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• v.24 no.4
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• pp.436-443
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• 2007

Anaerobic reductive dehalogenation of perchloroethene (PCE) was studied with lactate as the electron donor in a continuously stirred tank reactor (CSTR) inoculated with a mixed culture previously shown to dehalogenate vinyl chloride (VC). cis-1,2- dichloroethene (cDCE) was the dominant intermediate at relatively long cell retention times (>56 days) and the electron acceptor to electron donor molar ratio (PCE:lactate) of 1:2. cDCE was transformed to VC completely at the PCE to lactate molar ratio of 1:4, and the final products of PCE dehalogenation were VC (80%) and ethene (20%). VC dehalogenation was inhibited by cDCE dehalogenation. Propionate produced from the fermentation of lactate might be used as electron donor for the dehalogenation. Batch experiments were performed to evaluate the effects of increased hydrogen, VC, and trichloroethene (TCE) on VC dehalogenation which is the rate-limiting step in PCE dehalogenation The addition of TCE increased the VC dehalogenaiton rate more than an increase in the $H_2$ concentration, which suggests that the introduction of TCE induces the production of an enzyme that can comtabolize VC.

• Journal of Microbiology and Biotechnology
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• v.9 no.5
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• pp.576-581
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• 1999

Effects of signal sequences, protein sizes and dissolved oxygen on the secretion of human lysozyme from a recombinant yeast were experimentally characterized. The systems consisted of Saccharomyces cerevisiae host SEY2102 that was transformed with two different plasmids. These plasmids were identical with an exception to the plasmid pMC614, which contained the native yeast MFα1 sequence and the plasmid pMC632 with the non-native rat α-amylase signal sequence. The expression of human lysozyme was controlled by the ADHI promoter. The native yeast MFαl signal sequence was more efficient than the non-native rat α-amylase signal sequence in directing the secretion of human lysozyme. Lysozyme secreted with the α-amylase signal was retained inside the cells and released to the medium very slowly, thereby causing a lower cell growth rate and a decreased product secretion rate. Lysozyme was secreted more efficiently than invertase, which is an order of magnitude bigger in molecular size compared to lysozyme, which was under the direction of the MFαl signal sequence, suggesting that protein sizes may affect the secretion efficiency. When expressed in anaerobic conditions in the medium where the ADHI promoter was derepressed, the amount of lysozyme secreted was about twice higher than that of the aerobic culture. However, the secretion rates were identical. This result showed that the dissolved oxygen level may affect the efficiency of protein secretion only, and not the secretion rate of the product protein.

• KSBB Journal
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• v.21 no.4
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• pp.267-272
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• 2006

Five strains producing ethanol were isolated from soil near traditional alcohol production factory in Gwangju, Korea. One of the isolated strains maintained relatively stable ethanol production in shaking culture. The isolated strain KJ was proved to be Saccharomyces italicus, based on several biochemical and morphological tests containing assimilation of carbon compounds. In investment of the most suitable carbon for ethanol production, ethanol concentration of 5.46 g/L and yield of 0.53 g-ethanol/g-glucose were obtained in condition of glucose 10 g/L in YM medium. Experimental optimal conditions for ethanol fermentation by S. italicus KJ were as follows; temperature $30^{\circ}C$, initial pH 5.0, initial concentration 10% of glucose, anaerobic condition in the liquid cultivation. When enzymatically saccharified food wastes(SFW) were used as the production medium, ethanol production yield was 0.57 g-ethanol/g-reducing sugar. Therefore, SFW will contribute to lower the production cost of ethanol for industrial application.

• Korean Journal of Organic Agriculture
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• v.20 no.3
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• pp.327-343
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• 2012

In order to develop a high cellulolytic direct-fed microorganism (DFM) for ruminant productivity improvement, this study isolated cellulolytic bacteria from the rumen of Holstein dairy cows, and compared their cellulolytic abilities via DM degradability, gas production and cellulolytic enzyme activities. Twenty six bacteria were isolated from colonies grown in Dehority's artificial (DA) medium with 2% agar and cultured in DA medium containing filter paper at $39^{\circ}C$ for 24h. 16s rDNA gene sequencing of four strains from isolated bacteria showed that H8, H20 and H25 strains identified as Ruminococcus flavefaciens, and H23 strain identified as Fibrobacter succinogenes. H20 strain had higher degradability of filter paper compared with others during the incubation. H8 (R. flavefaciens), H20 (R. flavefaciens), H23 (F. succinogenes), H25 (R. flavefaciens) and RF (R. flavefaciens sijpesteijn, ATCC 19208) were cultured in DA medium with filter paper as a single carbon source for 0, 1, 2, 3, 4 and 6 days without shaking at $39^{\circ}C$, respectively. Dry matter degradability rates of H20, H23 and H25 were relatively higher than those of H8 and RF since 2 d incubation. The cumulative gas production of isolated cellulolytic bacteria increased with incubation time. At every incubation time, the gas production was highest in H20 strain. The activities of carboxymethylcellulase (CMCase) and Avicelase in the culture supernatant were significantly higher in H20 strain compared with others at every incubation time (p<0.05). Therefore, although further researches are required, the present results suggest that H20 strain could be a candidate of DFM in animal feed due to high cellulolytic ability.