• Journal of Microbiology and Biotechnology
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• v.29 no.12
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• pp.1882-1893
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• 2019

β-Glucosidases and β-xylosidases are two categories of enzymes that could cleave out non-reducing, terminal β-D-glucosyl and β-D-xylosyl residues with release of D-glucose and D-xylose, respectively. In this paper, two functional β-glucosidase Dth3 and β-xylosidase Xln-DT from Dictyoglomus thermophilum were heterologously expressed in E.coli BL21 (DE3). Dth3 and Xln-DT were relatively stable at 75℃ and were tolerant or even stimulated by glucose and xylose. Dth3 was highly tolerant to glucose with a K_i value of approximately 3 M. Meanwhile, it was not affected by xylose in high concentration. The activity of Xln-DT was stimulated 2.13-fold by 1 M glucose and 1.29-fold by 0.3 M xylose, respectively. Furthermore, the βglucosidase Dth3 and β-xylosidase Xln-DT showed excellent selectivity to cleave the outer C-6 and C-3 sugar moieties of ASI, which established an effective and green method to produce the more pharmacologically active CAG, an exclusive telomerase activator. We measured temperature, pH and dosage of enzyme using a single-factor experiment in ASI biotransformation. After optimization, the optimal reaction conditions were as follows: 75℃, pH 5.5, 1 U of Dth3 and 0.2 U of Xln-DT, respectively. Under the optimized conditions, 1 g/l ASI was transformed into 0.63 g/l CAG with a corresponding molar conversion of 94.5% within 3 h. This is the first report to use the purified thermostable and sugar-tolerant enzymes from Dictyoglomus thermophilum to hydrolyze ASI synergistically, which provides a specific, environment-friendly and cost-effective way to produce CAG.

• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1547-1556
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• 2009

A novel thermostable $\alpha$-glucosidase (TtGluA) from Thermoanaerobacter tengcongensis MB4 was successfully expressed in E. coli and characterized. The TtgluA gene contained 2,253 bp, which encodes 750 amino acids. The native TtGluA was a trimer with monomer molecular mass of 89 kDa shown by SDS-PAGE. The purified recombinant enzyme showed hydrolytic activity on maltooligosaccharides, p-nitrophenyl-$\alpha$-D-glucopyranide, and dextrin with an exotype cleavage manner. TtGluA showed preference for short-chain maltooligosaccharides and the highest specific activity for maltose of 3.26 units/mg. Maximal activity was observed at $60^{\circ}C$ and pH 5.5. The half-life was 2 h at $60^{\circ}C$. The enzyme showed good tolerance to urea and SDS but was inhibited by Tris. When maltose with the concentration over 50 mM was used as substrate, TtGluA was also capable of catalyzing transglycosylation to produce $\alpha$-1,4-linked maltotriose and $\alpha$-1,6-linked isomaltooligosaccharides. More importantly, TtGluA showed exclusive regiospecificity with high yield to produce $\alpha$-1,6-linked isomaltooligosaccharides when the reaction time extended to more than 10 h.

• Microbiology and Biotechnology Letters
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• v.41 no.2
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• pp.242-248
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• 2013

Endotoxins are part of the outer membrane of the cell wall of gram-negative bacteria and are continuously released during bacterial growth. Endotoxins typically induce severe sepsis and septic shock, which cause more than 50% of mortalities. Endotoxins are easily measured in the serum by the limulus amebocyte lysate (LAL) test. However, a nonspecific result is obtained, because the high concentration of serum proteins disturbs the enzyme reaction of the LAL test. In order to solve this problem, the LAL test was performed in this study after the centrifugation of the boiled serum samples to remove the impurities. As a result, among the various conditions examined, endotoxin measurement with the LAL test was the most accurate and repeatable after centrifugation of the boiled serum at $100^{\circ}C$. Moreover, the endotoxin was accurately and repeatedly measured from the prepared sera of mice that had been administered an intraperitoneal injection of purified lipopolysaccharides (LPS) or E. coli. Therefore, the application of centrifugation to remove impurities from boiled serum gives an accurate measurement of endotoxins in the sera of normal subjects or patients, and this will lead to the improved diagnosis and prevention of diseases caused by endotoxins. In addition, the centrifugation of boiled serum samples should be considered and included in the development of endotoxin test kits.

• Magazine of the Korean Society of Agricultural Engineers
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• v.45 no.6
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• pp.194-206
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• 2003

A pilot study was performed to examine the feasibility of the pond system for further polishing of treatment wetland effluent to agricultural reuse of reclaimed water. The constructed wetland and pond system was installed in Konkuk University and the effluent from septic tank of school building was used as an influent to the wetland system. The effluent of the wetland was used as an influent to pond systems. The influent concentrations of total coliform(TC), fecal coliform (FC), and E. coli were about $10^5$MPN/100 ml, and they were reduced to less than 10,000 MPN/100 ml on average after wetland treatments, showing over 95 % removal. And they were further reduced to less than 1,000 MPN/100 ml in average, showing over 85∼93 % removal after pond treatment. Turbidity and SS were improved effectively on average and their pond effluent concentration was about 4.5 NTU and 9.8 mg/L in average, respectively Average $BOD^5$ concentrations were also reduced substantially to 9.3 mg/L with about 83 % removal rate after wetland and pond treatment systems. Nutrients removal was relatively low and removal rate for T-N and T-P was less than 43 and 44%, respectively after wetland and pond treatment. Considering stable performance and effective removal of bacterial indicators as well as other water quality parameters, low maintenance, and cost-effectiveness, pond system was thought to be an effective and feasible alternative for agricultural reuse of reclaimed water. This paper describes a preliminary result Iron pilot study and further investigations are recommended on the optimum design parameters before full scale application.

• Journal of Biosystems Engineering
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• v.39 no.4
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• pp.345-351
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• 2014

Purpose: Rapid detection of bacteria is very important in agricultural and food industries to prevent many foodborne illnesses. The objective of this study was to develop a portable nuclear magnetic resonance (NMR)-based system to detect foodborne pathogens (E. coli). This study was focused on developing a method to detect low concentrations of magnetic nanoparticles using NMR techniques. Methods: NMR relaxometry was performed to examine the NMR properties of iron nanoparticle mixtures with different concentrations by using a 1 T permanent magnet magnetic resonance imaging system. Exponential curve fitting (ECF) and inverse Laplace transform (ILT) methods were used to estimate the NMR relaxation time constants, $T_1$ and $T_2$, of guar gum solutions with different iron nanoparticle concentrations (0, $10^{-3}$, $10^{-4}$, $10^{-5}$, $10^{-6}$, and $10^{-7}M$). Results: The ECF and ILT methods did not show much difference in these values. Analysis of the NMR relaxation data showed that the ILT method is comparable to the classical ECF method and is more sensitive to the presence of iron nanoparticles. This study also showed that the spin-spin relaxation time constants acquired by a Carr-Purcell-Meiboom-Gill (CPMG) pulse sequence are more useful for determining the concentration of iron nanoparticle solutions comparwith the spin-lattice relaxation time constants acquired by an inversion recovery pulse sequence. Conclusions: We conclude that NMR relaxometry that utilizes CPMG pulse sequence and ILT analysis is more suitable for detecting foodborne pathogens bound to magnetic nanoparticles in agricultural and food products than using inversion recovery pulse sequence and ECF analysis.

• Asian-Australasian Journal of Animal Sciences
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• v.13 no.11
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• pp.1554-1560
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• 2000

Ninety-six crossbred (Large White${\times}$Landrace${\times}$Duroc) pigs, weaned at 35 days of age, were assigned to four dietary treatments in order to investigate the effects of oral antibiotics on the performance and the intestinal microflora of weanling pigs. Pigs were fed either a basal diet, without antibiotics, or the basal diet plus either 50 ppm acetylspiramycin, 50 ppm olaquindox, or 100 ppm bacitracin zinc. The pigs were housed eight per pen with three pens per treatment in an environmentally controlled nursery. Ten days after weaning, three pigs from each treatment were slaughtered and intestinal pH, microflora, and volatile fatty acid concentration were determined. At the end of the 4 week trial, the remaining pigs were weighed and feed consumption was measured. Average daily gains for pigs fed acetylspiramycin, olaquindox, bacitracin zinc and the control diet were 0.43, 0.40, 0.37, and 0.34 kg per day (p=0.001), respectively. Antibiotic addition did not modify feed intake, but acetylspiramycin improved feed conversion (p=0.003). In comparison with the control, acetylspiramycin significantly increased Bifidobacteria numbers in the jejunum (p=0.082) and ileum (p=0.014) and decreased total bacterial counts throughout the intestine (p<0.01 except for the ileum where p=0.079). Acetate production was significantly lower in the cecum (p=0.028) and colon (p=0.079) of pigs fed acetylspiramycin. In addition to increasing numbers of Bifidobacteria in the jejunum (p=0.082) and ileum (p=0.014), olaquindox increased Lactobacillus in the jejunum (p=0.004) and decreased E. coli in the colon (p=0.022). Bacitracin zinc increased Lactobacillus numbers in the jejunum (p=0.004) and Bifidobacterium concentrations in the jejunum (p=0.082) and ileum (p=0.014).

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.7
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• pp.1015-1020
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• 2012

An experiment was conducted to evaluate the effects of bacteriophage supplementation on egg performance, egg quality, excreta microflora, and moisture content in laying hens. A total of 288 Hy-line brown commercial laying hens (36-wk-old) were randomly allotted to 4 treatments in this 6-wk trial and dietary treatments included: i) CON, basal diet; ii) T1, CON+0.020% bacteriophage; iii) T2, CON+0.035% bacteriophage; iv) T3, CON+0.050% bacteriophage. There were 6 replicates for each treatment with 6 adjacent cages (2 hens/cage). Laying hens in T2 and T3 treatments had higher (p<0.05) egg production than those in CON and T1 treatments during wk 0 to 3. In addition, egg production in T1, T2, and T3 treatments was increased (p<0.05) compared with that in CON treatment during wk 4 to 6. At wk 4 and 5, birds in T2 group had higher (p<0.05) HU than those in CON. In addition, at wk 5 and 6, HU in birds fed T1 and T3 diets was greater (p<0.05) than those fed CON diet. E. coli and Salmonella spp. concentrations in excreta were decreased (p<0.05) by T1, T2, and T3 treatments. However, egg weight, egg shell color, yolk height, yolk color unit, egg shell strength, egg shell thickness, egg gravity, and excreta moisture content were not influenced by dietary treatments during the entire experimental period. In conclusion, bacteriophage supplementation has beneficial effects on egg production, egg albumen, and excreta microflora concentration in laying hens.

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.5
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• pp.674-681
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• 2012

Two experiments were conducted to investigate the effects of supplemental glutamine on growth performance, plasma parameters and LPS-induced immune response of weaned barrows after castration. In experiment 1, forty-eight weaned male piglets were used and fed maize and soybean meal diets supplemented with 0 (Control) or 2% L-Gln (Gln+) for 25 days. The results indicated that the Gln+ group tended to increase average daily gain compared to control in stages of days 7 to 14 and 0 to 25. The Gln+ had significantly better feed efficiency than the control group did during days 14 to 25 and 0 to 25. The plasma blood urea nitrogen and alkaline phosphatase contents of Gln+ group were higher than those of the control group on day 14 post-weaning. In experiment 2, sixteen weaned male piglets were injected with E. coli K88+ lipopolysaccharide (LPS) on day 14 post-weaning. The results showed that the Gln+ group had lower concentrations of plasma adrenocorticotrophic hormone and cortisol than the control group on day 14 pre-LPS challenge. In addition, Gln+ group had higher plasma IgG concentration than the control group for pre- or post-LPS challenged on day 14 post-weaning. In summary, dietary supplementation of Gln was able to alleviate the stressful condition and inflammation associated with castration in weaned barrows, and to improve their immunity and growth performance in the early starter stage.

• Microbiology and Biotechnology Letters
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• v.14 no.6
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• pp.455-460
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• 1986

Dane particle was prepared from the plasma of chronic HBsAg carrier with high levels of HBsAg activity. DNA extracted front Dane particle core after a DNA polymerase reaction with $\alpha$-($^{32}$P) dNTP, was identified as HBV DNA by liquid scintillation counter and agarose gel electrophoresis-G.M. counting. To produce Hepatitis B surface antigen for use as a vaccine against Hepatitis B virus infection, yeast strains harboring recombinant plasmid with Apase promoter was used. Recombinant plasmid was construced from pHBV 130 and pAN 82, transformed into E coli, and then transferred into yeast strains. HBsAg was produced by derepression in Burkholder minimal medium with controlled inorganic phosphate concentration. The kinetics of HBsAg production was also investigated. Total HBsAg activity increased rapidly between 3 and 6 hours after transfer to phosphate-free medium and reached a maximum at around 9th hour. The transfer into phosphate-free medium after 6 hours in high phosphate cell growth medium gave maximum activity.

• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.659-665
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• 2016

The oligosaccharides in human milk constitute a major innate immunological mechanism by which breastfed infants gain protection against infectious diarrhea. Clostridium difficile is the most important cause of nosocomial diarrhea, and the C-terminus of toxin A with its carbohydrate binding site, TcdA-f2, demonstrates specific abolishment of cytotoxicity and receptor binding activity upon diethylpyrocarbonate modification of the histidine residues in TcdA. TcdA-f2 was cloned and expressed in E. coli BL21 (DE3). A human milk oligosaccharide (HMO) mixture displayed binding with TcdA-f2 at 38.2 respond units (RU) at the concentration of 20 μg/ml, whereas the eight purified HMOs showed binding with the carbohydrate binding site of TcdA-f2 at 3.3 to 14 RU depending on their structures via a surface plasma resonance biosensor. Among them, Lacto-N-fucopentaose V (LNFPV) and Lacto-N-neohexaose (LNnH) demonstrated tight binding to TcdA-f2 with docking energy of −9.48 kcal/mol and −12.81 kcal/mol, respectively. It displayed numerous hydrogen bonding and hydrophobic interactions with amino acid residues of TcdA-f2.