• Korean Journal of Veterinary Research
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• v.43 no.2
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• pp.255-261
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• 2003

The antigenicity of nonspecific immunostimulator BARODON$^{(R)}$, a newly developed drug, was investigated by tests for passive cutaneous anaphylaxis (PCA) and active systemic anaphylaxis (ASA) in mice and guinea pigs. In ASA test using guinea pigs, there were no significant clinical symptoms in all individuals of low(0.3%) and high(3%) dose of both groups treated with only BARODON$^{(R)}$ and cotreated with BARODON$^{(R)}$ and adjuvant group. In PCA test, blue spots of Evan's were observed from $2^6$ to $2^{10}$ in homologous group and from $2^2{\sim}2^5$ dilution rate in heterologous group of BSA treated positive control group. However, intradermal sensitization with antiserum obtained from low (0.3%) and high (3%) dose of BARODON$^{(R)}$ only treatment group and treated-with-adjuvant group, followed by intravenous injection of respective antigen and Evan's blue mixture (1:1) showed no blue spot observed. In conclusion, BARODON$^{(R)}$, as showed in ASA and PCA test, did not cause anaphylatic shock when treated 3 and 10 times higher than clinically intended dose, nor induce IgE, so that might not have antigenic properties in mice and guinea pigs.

• Resources Recycling
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• v.28 no.6
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• pp.96-105
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• 2019

Automobile shredder residue (ASR) is the final waste produced when end-of-life vehicles (ELVs) are shredded. ASR can be separated using mineral-processing operations such as comminution, air classification, magnetic separation, and/or electrostatic separation. In this work, trajectory analyses of conductors (copper) and non-conductors (glass) in the ASR have been carried out using induction electrostatic separator for predicting or improving the ASR-separation efficiency. From results of trajectory analysis for conductors, the trajectories of copper wire by observation versus simulation for coarse particles of 0.5 and 0.25 mm showed consistent congruity. The observed 0.06 mm fine-particles trajectory was deflected toward the (-) attractive electrode owing to the charge-density effects due to the particle characteristics and relative humidity. In the case of non-conductors, the actual trajectory of dielectric glass deflected toward the (-) electrode, showing characteristics similar to those of conductive particles. The analyses of stereoscopic microscope and SEM & EDS found heterologous materials (fine ferrous particles and conductive organics) on the glass surface. This demonstrates the glass decreasing separation efficiency for non-ferrous metals during electrostatic separation for the recycling of ASR. Future work will require a pretreatment process for eliminating impurities from the glass and advanced trajectory-simulation processes.

• Microbiology and Biotechnology Letters
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• v.33 no.4
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• pp.274-280
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• 2005

Pasteurella multocida is known to cause widespread infections in husbandry. To induce homologous and heterologous immunity against the infections, outer membrane proteins (OMPs) in the envelope of P. multocida are thought to be attractive vaccine candidates. Outer membrane protein H is considered as the major component of OMPs. In this study, a gene for OmpH was isolated from pathogenic P. multocida serogroup A. The gene was composed of 1,047 nucleotides coding 348 amino acids with signal peptide of 20 amino acids. The amino acid composition showed about 80 to 98 per cent sequence homologies among other 10 strains of P. multocida serogroup A, reported so far. A recombinant ompH, from which signal peptide was truncated, was generated using pRSET A to name 'pRSET A/OmpH-F2'. The pRSET A/OmpH-F2 was well expressed in E. coli BL21(DE3). The truncated OmpH was purified using nickel-nitrilotriacetic acid (Ni-NTA) affinity column chromatography. Its molecular weight was registered to be 40 kDa on SDS-PAGE gel. In order to generate immunesera against the OmpH, 50 ug of the protein was intraperitoneally injected into mice three times. The anti-OmpH immuneserum recognized about $5{\times}10^{-2}$ng quantity of the purified OmpH. It can be used for an effective vaccine production to prevent fowl cholera caused by pathogenic P. multocida (Serogroup A).

• Microbiology and Biotechnology Letters
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• v.32 no.3
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• pp.199-205
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• 2004

The nucleotide sequence of 8.6-kb EcoRI fragment containing sucrose phosphorylase gene isolated from Bifidobacterium longum SJ32 was determined. It was found that the fragment contained five open reading frames including the gene cluster for sucrose utilization such as a sucrose phosphorylase (ScrP), a sucrose transporter (ScrT), and a GalR-LacI-type transcriptional regulator (ScrR) identified by amino acid homology. Each gene showed over 94% amino acid homology among various B. longum strains. Genomic organization of the gene cluster is the same as those of other strains of B. longum but different from that of B. lactis. In spite of high homology of each gene among B. longum strains, the difference of flanking sequences of the gene cluster between strains SJ32 and NCC2705 insinuates the horizontal transfer of scrPTR between B. longum strains. The increase of sucrose phosphorylase activity in heterologous E. coli system by the co-expression of scrT with scrP against the single expression of scrP was measured. It seems to be the result of sucrose uptake increment by scrT in the host and is an indirect evidence that scrT is the gene for sucrose transport. The existence of multiple sucrose uptake systems in B. longum is supposed from the findings of several genes besides scrPTR involved in sucrose uptake in the genome of B. longum NCC2705.

• Applied Biological Chemistry
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• v.48 no.4
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• pp.345-351
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• 2005

To access the natural products of uncultured microorganisms, we constructed and screened the metagenomic DNA libraries by using a cosmid vector and DNA inserts isolated directly from soil. Initial screening of the libraries in Escherichia coli resulted in the isolation of several clones that produce a dark brown color when grown in LB medium. One of the positive clones, designed pYS85C, was transposon mutagenized and the DNA surrounding the transposon insertions in cosmids that no longer conferred the production of brown pigment to E. coli was sequenced. Annotation of the pYS85C sequence obtained from the transposon mutagenesis experiment indicated a single 393 amino acid open reading frame (ORF) with a molecular mass of about 44.5 kDa, predicted to be a 4-hydroxyphenylpyruvate dioxygenases (HPPDs), was responsible for the observed brown pigment. In a BLAST search against deposited sequence, the translated protein from this ORF showed moderate-level identity (>60%) to the other known HPPDs and was most conserved in the C-terminal region of the protein. These results show that genes involved in natural product synthesis can be cloned directly from soil DNA and expressed in a heterologous host, supporting the idea that this technology has the potential to provide novel natural products from the wealth of environmental microbial diversity and is a potentially important new tool for drug discovery.

• Journal of Applied Biological Chemistry
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• v.58 no.4
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• pp.325-332
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• 2015

Rapeseed (Brassica napus) is now the second largest oilseed crop after soybean. Cold temperature tolerance is an important agronomic trait in winter rapeseed that determines the plant's ability to control below freezing temperatures. To improve cold tolerance of rapeseed plants, an expression vector containing an Barley Ice recrystallization inhibition protein (HvIRIP) cDNA driven by a cauliflower mosaic virus 35S promoter was transferred into rapeseed plants. Transgenic expression of HvIRIP was proved by southern- and northern-blot analyses. The level of freezing tolerance of transgenic $T_3$ plants was found to be significantly greater than that of wild-type rapeseed plants by freezing assay. Proline accumulation during cold stress was also highly induced in the transgenic rapeseed plants. The transgenic plants exhibited considerable tolerance against oxidative damage induced by cold stress. Our results indicated that heterologous HvIRIP expression in transgenic rapeseed plants may induce several oxidative-stress responsive genes to protect from cold stress.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.1-7
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• 1989

To enhance the capability of starch fermentation of the transformant TSD-14, the heat treated protoplasts of TSD-14 were fused with the protoplasts of C. tropicalis (lys$^-$) in the presence of 30% (w/ v) PEG and 20 mM CaC1$_2$. Fusants were selected by nutritional complementation on minium medium and the fusion frequency was 4.4$\times$10$^{-5}$. All fusants tested were possessed of complemented traits concerning carbon compound assimilation, and the cell volumes of the fusants were approximately 1.5 times larger than the parental strains. The fusants were genetically very stable, and were able to hydrolyze alpha 1,4-glucosidic linkage as well as alpha 1,6-linkage of starch contrary to one of parents TSD-14, The most promising fusant FSC-14-75 produced 8.7% (v/v) of ethanol from 15% liquefied potato starch medium, but the result was enhanced to 9.3% (v/v) by addition of 0.3% peptone. The corresponding fermentation efficiency was 86.0%.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.14-18
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• 1989

As the final experiment to assess the possibility of industrial application of FSC14-75, ethanol productivity from liquefied sweet potato starch was examined in a pilot scale of 300 liters. FSC14-75 produced 6.6％(v/v) of ethanol from 13.3％ of liquefied sweet potato starch in 8 days, and the residual sugar was 3.15％. The corresponding efficiency was 70％ of the theoretical maximum. Since we could isolate unicellular cell and flocculent cell from the fermentation broth, we designated them FSC14-75(S) and FSC14-75(F), respectively. We investigated ethanol productivity of FSC14-75(F) compared with that of FSC14-75(S) from liquefied potato starch in a mini·tar tormentor scale of 2.5 liters. FSC14-75(F) was found more favorable than the counterpart in terms of ethanol productivity, and produced 8.1％(v/v) of ethanol from 15％ of liquefied potato starch with an efficiency of 75％. In a pilot scale fermentation with 15％ of liquefied sweet potato starch, ethanol productivity of FSC14-75(F) reached maximum level of 7.7％(v/v) after 8 days, and the residual sugar was 1.9％. However, the ethanol productivity was not enhanced by a supplementary addition of Thermamyl to the fermentation broth after sterilization.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.8-13
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• 1989

The optimum conditions for ethanol fermentation and ethanol productivity of the fusant ESC-14-15 were examined in a mini-jar formentor scale (working volume : 2.5 liters) to assess the possibility of practical application. Addition of yeast extract to fermentation broth greatly enhanced the ethanol productivity and shortened the period of fermentation. The pH 4.2 was more favorable than pH 5.5 with respect to ethanol productivity and fermentation speed. The optimum concentration of liquefied potato starch for ethanol fermentation of FSC-14-15 was 15%(w/v) and the corresponding productivity was 8.7%(v/v) of ethanol with an efficiency of 80.6% to the theoretical maximum. When the fresh fermentation broth containing 20% of liquefied potato starch was inoculated with love(v/v) of inoculum, the fusant FSC-14-75 produced 11.0%(v/v) of ethanol in 4 days, which is considered comparable to that from an industrial process. From the liquefied cassava starch or the equal mixture of liquefied barley and sweet potato starch prepared according to the same method as in the industrial process except saccharification step, the fusnnt FSC-14-75 produced 8.5%(v/v) or 7.6%(v/v) of ethanol in 4 days, respectively.

• Journal of Microbiology and Biotechnology
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• v.18 no.8
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• pp.1386-1392
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• 2008

A gene (sll0158) putatively encoding a glycogen branching enzyme (GBE, E.C. 2.4.1.18) was cloned from Synechocystis sp. PCC6803, and the recombinant protein expressed and characterized. The PCR-amplified putative GBE gene was ligated into a pET-21a plasmid vector harboring a T7 promoter, and the recombinant DNA transformed into a host cell, E. coli BL21(DE3). The IPTG-induced enzymes were then extracted and purified using Ni-NTA affinity chromatography. The putative GBE gene was found to be composed of 2,310 nucleotides and encoded 770 amino acids, corresponding to approx. 90.7 kDa, as confirmed by SDS-PAGE and MALDI-TOF-MS analyses. The optimal conditions for GBE activity were investigated by measuring the absorbance change in iodine affinity, and shown to be pH 8.0 and $30^{\circ}C$ in a 50 mM glycine-NaOH buffer. The action pattern of the GBE on amylose, an $\alpha$-(1,4)-linked linear glucan, was analyzed using high-performance anion-exchange chromatography (HPAEC) after isoamylolysis. As a result, the GBE displayed $\alpha$-glucosyl transferring activity by cleaving the $\alpha$-(1,4)-linkages and transferring the cleaved maltoglycosyl moiety to form new $\alpha$-(1,6)-branch linkages. A time-course study of the GBE reaction was carried out with biosynthetic amylose (BSAM; $M_p{\cong}$8,000), and the changes in the branch-chain length distribution were evaluated. When increasing the reaction time up to 48 h, the weight- and number-average DP ($DP_w$ and $DP_n$) decreased from 19.6 to 8.7 and from 17.6 to 7.8, respectively. The molecular size ($M_p$, peak $M_w{\cong}2.45-2.75{\times}10^5$) of the GBE-reacted product from BSAM reached the size of amylose (AM) in botanical starch, yet the product was highly soluble and stable in water, unlike AM molecules. Thus, GBE-generated products can provide new food and non-food applications, owing to their unique physical properties.