• Journal of Microbiology and Biotechnology
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• v.25 no.7
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• pp.999-1006
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• 2015

The endo-polygalacturonase gene (endo-pgaA) was cloned from DNA of Aspergillus niger SC323 using the cDNA synthesized by overlapping PCR, and successfully expressed in Saccharomyces cerevisiae EBY100 through fusing the α-factor signal peptide of yeast. The fulllength cDNA consists of 1,113 bp and encodes a protein of 370 amino acids with a calculated molecular mass of 38.8 kDa. After induction by galactose for 48 h, the activity of recombinant endo-PgaA in the culture supernatant can reach up to 1,448.48 U/mg. The recombinant protein was purified to homogeneity by ammonium sulfate precipitation and gel filtration column chromatography and subsequently characterized. The optimal pH and temperature of the purified recombinant enzyme were 5.0 and 50℃, respectively. The Michaelis-Menten constant (K_m) and maximal velocity (V_max) of the enzyme for pectin were 88.54 μmol/ml and 175.44 μmol/mg/min, respectively. The enzyme activity was enhanced by Ca²⁺, Cu²⁺, and Na⁺, and strongly inhibited by Pb²⁺ and Mn²⁺. The pectin hydrolysates were mainly galacturonic acid and other oligo-galacturonates. Therefore, these characteristics suggest that the recombinant endo-PgaA may be of potential use in the food and feed industries.

• Microbiology and Biotechnology Letters
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• v.46 no.2
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• pp.127-134
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• 2018

In this study, 79 bacterial isolates were collected from the surface of marine algae Umbraulva japonica. As a result of analysis of 16s rRNA gene sequence, the 79 isolated bacteria were divided into 4 major groups: [Proteobacteria (74.69%), Actinobacteria (2.53%), Fimicutes (2.53%), and Bacteroidetes (20.25%)] - 7 classes (Actinobacteria, Flavobacteria, Sphingobacteria, Baciili, Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria), 12 orders, 17 families and 31 genera. The newly isolated 3 strains could be novel species because of less than 97% similarity in 16s rRNA sequence. Therefore, it is considered that additional experiments should be conducted together with the standard strain. Analysis of 79 bacterial antibacterial activity against human and fish pathogens, such as Edwardsiella tarda, Vibrio harveyi, Streptococcus iniae, Steptococcus parauberis, Escherichia coli, Steptococcus mutans, Listeria monocytogenes and Vibrio vulnificus, was performed by using the supernatant liquid and pellet. As a result, pellet of UJT9, UJT20 and UJR17 showed antibacterial activity against V. vulnificus, UJR17 also showed antibacterial activity against S. parauberis. UJT7 and UJT20, UJR17 have been identified as Bacillus sp. and Pseudomonas sp. and it may be safely assented that it's beneficial to carry out additional experiments for various applications.

• Microbiology and Biotechnology Letters
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• v.27 no.5
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• pp.419-425
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• 1999

Prophylactic effects of Bifidobacterium longum HY8001, Korean isolate, against Escherichia coli O157:H7 and Salmonella typhimurium DT104 enteric infection were examined at four groups of specific pathogen free(SPF)-ICR mouse for each pathogen. B. longum HY8001+B. typhimurium DT104+B. longum HY8001(BL+ST+BL) group and B. longum HY8001+E. coli O157:H7+B. longum HY8001(BL+E+BL) group were fed with B. longum HY8001 before and after E. coli O157:H7 or s. typhimurium DT104 challenge, while B. longum HY8001+S. typhimurium DT104(BL+ST) and B. longum HY8001+e. coli O157:H7(BL+E) groups were fed with B. longum HY8001 only before E. coli O157:H7 or S. typhimurium DT104 challenge. E. coli O157:H7(E) and S. typhimurium DT104(ST) groups were challenged with each pathogen without B. longum HY8001 administration and control groups were administered with phosphate buffered solution(PBS). After the oral administration with B. longum HY8001(109cfu), th emice were challenged with E. coli O157:H7(2$\times$1010cfu) or S. typhimurium DT104(108cfu) and the mortality rate and the fecal shedding of challenged pathogen were also examined define the reactivity of the B. longum HY8001. Production of toxin neutralizing substance(s) of B. longum HY8001 was determined by cell cytotoxicity assay using Vero cells. Fecal shedding of th eS. typhimurium DT104 was significantly decreased in BL+ST+BL group fed with B. longum HY8--1 before and after challenge(p<0.05), while the fecal shedding s of S. typhimurium DT104 in BL+ST and St groups remained more than 106cfu. the protective effect of the B. longum HY8001 against E. coli O157:H7 was significantly high only in BL+E+BL group fed with b. longum Hy8001 before and after E. coli O157:H7 challenge from the result of fecal E. coli O157:H7 isolation rate, mortality rate, and intestinal contents culture to detect E. coli O157:H7. the mortality rate of the BL+e and E groups. The cytopathic effect (CPE) of the Vero cytotoxin (Shiga like toxin I & II) in Vero cell was neutralized in B. longum HY8001 culture supernatant added wells which indicate the presence of soluble Vero cytotxin neutralizing substance(s) in B. longum HY8001 culture suprnatant.

• Microbiology and Biotechnology Letters
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• v.36 no.2
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• pp.121-127
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• 2008

A strain IDCC 9203 isolated from infant feces was identified as Lactobacillus johnsonii on the basis of 16S rDNA sequence analysis. L. johnsonii IDCC 9203 was highly resistant to acid (MRS broth at pH 2.3) and bile (MRS broth with 0.3% oxgall). The antibacterial activities of L. johnsonii IDCC 9203 was examined against Salmonella typhimurium KCTC 2054. The growth of S. typhimurium KCTC 2054 was inhibited by the cell-free culture supernatant (at pH 4.0) of L. johnsonii IDCC 9203 as well as by the respective control (MRS broth at pH 4.0). Antimicrobial effect against S. typhimurium KCTC 2054 of L. johnsonii IDCC 9203 was probably due to the lactic acid. By an in vitro cell adhesion model, L. johnsonii IDCC 9203 preincubated or coincubated with Caco-2 cells reduced the adhesion of S. typhimurium KCTC 2054 to Caco-2 cells by 74% or 47.1%, respectively. Also in an in vivo model, L. johnsonii IDCC 9203 was colonized in mice intestines which were disrupted by ampicillin treatment. Its proliferation in the mice intestines reduced abnormal salmonella growth from $10^9CFU/g$ feces to $10^5CFU/g$ feces as an indigenous level. The results obtained in this study suggest that L. johnsonii IDCC 9203 may be a potential probiotic strain.

• Journal of Microbiology and Biotechnology
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• v.21 no.10
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• pp.1012-1020
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• 2011

A gene coding for an endoglucanase (EglA), of the glycosyl hydrolase family 12 and derived from Aspergillus niger VTCC-F021, was cloned and sequenced. The cDNA sequence, 717 bp, and its putative endoglucanase, a 238 aa protein with a predicted molecular mass of 26 kDa and a pI of 4.35, exhibited 98.3-98.7% and 98.3-98.6% identities, respectively, with cDNA sequences and their corresponding endoglucanases from Aspergillus niger strains from the GenBank. The cDNA was overexpressed in Pichia pastoris GS115 under the control of an AOX1 promoter with a level of 1.59 U/ml culture supernatant, after 72 h of growth in a YP medium induced with 1% (v/v) of methanol. The molecular mass of the purified EglA, determined by SDS-PAGE, was 33 kDa, with a specific activity of 100.16 and 19.91 U/mg toward 1% (w/v) of ${\beta}$-glucan and CMC, respectively. Optimal enzymatic activity was noted at a temperature of $55^{\circ}C$ and a pH of 5. The recombinant EglA (rEglA) was stable over a temperature range of $30-37^{\circ}C$ and at pH range of 3.5-4.5. Metal ions, detergents, and solvents tested indicated a slightly inhibitory effect on rEglA activity. Kinetic constants ($K_m$, $V_{max}$, $k_{cat}$, and $k_{cat}/K_m$) determined for rEglA with ${\beta}$-glucan as a substrate were 4.04 mg/ml, 102.04 U/mg, 2,040.82 $min^{-1}$, and 505.05, whereas they were 10.17 mg/ml, 28.99 U/mg, 571.71 $min^{-1}$, and 57.01 with CMC as a substrate, respectively. The results thus indicate that the rEglA obtained in this study is highly specific toward ${\beta}$-glucan. The biochemical properties of rEglA make it highly valuable for downstream biotechnological applications, including potential use as a feed enzyme.

• Journal of Microbiology and Biotechnology
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• v.23 no.11
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• pp.1519-1528
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• 2013

Bacillus pumilus SG2, a halotolerant strain, expresses two major chitinases designated ChiS and ChiL that were induced by chitin and secreted into the supernatant. The present work aimed to obtain a mutant with higher chitinolytic activity through mutagenesis of Bacillus pumilus SG2 using a combination of UV irradiation and nitrous acid treatment. Following mutagenesis and screening on chitin agar and subsequent formation of halos, the mutated strains were examined for degradation of chitin under different conditions. A mutant designated AV2-9 was selected owing to its higher chitinase activity. To search for possible mutations in the whole operon including ChiS and ChiL, the entire chitinase operon, including the intergenic region, promoter, and two areas corresponding to the ChiS and ChiL ORF, was suquenced. Nucleotide sequence analysis of the complete chitinase operon from the SG2 and AV2-9 strains showed the presence of a mutation in the catalytic domain (GH18) of chitinase (ChiL). The results demonstrated that a single base change had occurred in the ChiL sequence in AV2-9. The wild-type chitinase, ChiL, and the mutant (designated ChiLm) were cloned, expressed, and purified in E. coli. Both enzymes showed similar profiles of activity at different ranges of pH, NaCl concentration, and temperature, but the mutant enzyme showed approximately 30% higher catalytic activity under all the conditions tested. The results obtained in this study showed that the thermal stability of chitinase increased in the mutant strain. Bioinformatics analysis was performed to predict changes in the stability of proteins caused by mutation.

• Journal of Korean Society of Environmental Engineers
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• v.30 no.1
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• pp.31-36
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• 2008

This work was performed to evaluate the effect of membrane material and structure on fouling in a submerged membrane bioreactor(MBR). Three types of microfiltration membranes with the same pore size of 0.1 $\mu$m but different materials, polytetrafluoroethylene (PTFE), polycarbonate(PCTE) and polyester(PETE), were used. While PETE membrane exhibited the most rapid flux decline throughout the operation, PCTE and PTFE had a similar tendency with regard to permeability. Difference in permeability between PETE and the other membranes gradually decreased with time, which was probably due to chemical cleaning. The higher TOC rejection of PETE membrane could be attributable to its faster fouling, resulting from a larger amount of foulants to get attached to the membrane in a shorter time. DOC fractionation using a DAX-8 resin showed that the composition of each fraction between the supernatant and permeates did not change significantly with operation time, indicating that membrane hydrophilicity/hydrophobicity was not a dominant factor affecting to MBR fouling in this study. Compared to other membranes, the fouling of PETE membrane was more influenced by pore clogging (irreversible fouling), which would probably contribute to a higher organic rejection of the PETE membrane.

• The Korean Journal of Pesticide Science
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• v.20 no.2
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• pp.152-158
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• 2016

The cause of death was investigated with several dead cabbage moth larvae in breeding box. Bacterial strains were isolated and selected from the dead larvae by bioassay. One of them was identified as Serratia marcescens used by morphological characteristics and gene sequencing. S. marcescens were cultured by Luria Bertani (LB) media broth for bioassay. When 100-fold dilution of culture broth to third larvae of diamondback moth, Plutella xylostella, it was showed a 100% mortality at 2 days after treatment, and only 10-fold dilution of supernatant liquid was showed 86.6% mortality. When the culture broth of S. marcescens was applied to the larvae of beet armyworm, Spodoptera exigua, contact and feeding toxicity were 20 and 8% of mortality, respectively. Otherwise, when the culture broth of S. marcescens was applied to 5 major plant pathogens, antibacterial activities against Fusarium oxysporum, Rhizoctonia solani, Colletotrichum gloeosporioides, Phytophthora capsici and Sclerotinias clerotiorum were 4.7, 11.3, 20, 15.7 and 42.6%, respectively. Also, degradation ability of S. marcescens against protein and chitin were examined.

• Journal of Sasang Constitutional Medicine
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• v.16 no.1
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• pp.120-129
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• 2004

Yangkyuk-Sanhwa-Tang(YST) has been widely used as a formula for the Soyangin cerebral infarction (CI) patients according to Sasang constitutional philosophy. Brain cells produce cytokines and chemokines during the inflammatory process after stroke both in animal models and in patients. Previously, regulation of serum cytokine levels by YSThas been observed in individuals at the acute stage of CI disease, but there have not been other scientific investigations on YST. The author investigated the effect of YST on theproduction of various cytokines using peripheral blood mononuclear cells (PBMCs)from the Soyangin (CI) patients, and Soyangin normal group. The cytokine production was analyzed using enzyme-linked immunosorbent assay (ELISA). The amount of interleukin (IL)-1, IL-1, IL-6, IL-8, and tumor necrosis factor (TNF)- in culture supernatant significantly increased in the LPS-treated cells compared with unstimulated-cells (P < 0.05). However, in LPS-stimulated PBMCs, cytokines level in CI patients group was higher than that of normal group. YST (1 mg/ml) significantly inhibited IL-1, IL-1, and IL-8 production in PBMCs stimulated with LPS (about 85% for IL-1, 87% for IL-1, and 53% for IL-8, P < 0.05), but did not significantly inhibit IL-6 and TNF- production in the CI patients group. We also show that YST significantly increased LPS-induced IL-1, IL-6, and TNF- production in the normal group. Thesedata suggest that YST has a regulatoryeffect on the cytokine production, which might explain its beneficial effect in the treatment of CI.

• The Journal of Korean Obstetrics and Gynecology
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• v.27 no.2
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• pp.46-58
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• 2014

Objectives: This study was designed to investigate intestinal immune system activation and anti-metastatic effect on cancer cells by orally administered extracts of Boyanghwano-tang. Methods: To observe immunomodulating effects of Boyanghwano-tang on Peyer's patch cells, we measured cytokines GM-CSF, IL-4. In addition to observing effects of Boyanghwano-tang on hematopoiesis, we measured proliferation of bone marrow cells mediated by Peyer's patch cells in vitro. IgA induction activated in intestinal content and serum was measured to observe the effect of orally administered Boyanghwano-tang on mucosal immune system. After administering ovalbumin (OVA) with Boyanghwano-tang, Proliferation of Peyer's patch cell was measured to investigate gut immunostimulatory effect. Anti-metastatic experiments were conducted in vivo mouse model by using colon 26-M3.1 carcinoma cell. Results: The amounts of GM-CSF and IL-4 in the culture supernatant of Peyer's patch cells were significantly increased compared to the control group. The proliferation of bone marrow cell was significantly up-regualted with Boyanghwano-tang. These results indicate that oral administration of Boyanghwano-tang enhances the secretion of hematopoietic growth factors such as GM-CSF and IL-4 from Peyer's patch cells, and these cytokines also act on modulator of bone marrow cell proliferation. After orally administering OVA with Boyanghwano-tang, IgA induction and Proliferation of peyer's patch cell was up-regulated with Boyanghwano-tang. These results means orally administered Boyanghwano-tang activates intestinal immune system and has an inhibitory effect on tumor metastasis. In addition, We found that orally administered Boyanghwano-tang significantly inhibited tumor metastasis in vivo. Conclusions: Orally administered Boyanghwano-tang appears to have considerable activity on the anti-metastasis by activation of immune system.