• Journal of Microbiology and Biotechnology
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• v.20 no.1
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• pp.132-137
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• 2010

The skc gene encoding streptokinase (SK) with a molecular mass of approximately 47.4 kDa was cloned from Streptococcus equisimilis ATCC 9542 and heterologously overexpressed in Streptomyces lividans TK24 and E. coli using various strong promoters. When the promoter for sprT [Streptomyces griseus trypsin (SGT)] was used in the host S. lividans TK24, a 47.4-kDa protein was detected along with a smaller hydrolyzed protein (44 kDa), suggesting that posttranslational hydrolysis had occurred as has been reported in other expression systems. The casein/plasminogen plate assay revealed that the plasmid construct containing the SGT signal peptide was superior to that containing the SK signal peptide in terms of SK production. Maximal production of SK was calculated to be about 0.25 unit/ml of culture broth, a value that was five times higher than that obtained with other expression systems using ermE and tipA promoters in the same host. When the skc gene was expressed in E. coli BL21(${\Delta}DE3$)pLys under the control of the T7 promoter, a relatively large amount of SK was expressed in soluble form without hydrolysis. SK activity in E. coli/pET28a-$T7_pSK_m$ was more than 2 units/ml of culture broth, even though about half of the expressed protein formed an inactive inclusion body.

• Molecules and Cells
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• v.28 no.6
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• pp.553-558
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• 2009

In the periphery, a galectin-1 receptor, CD7, plays crucial roles in galectin-1-mediated apoptosis of activated T-cells as well as progression of T-lymphoma. Previously, we demonstrated that $NF-{\kappa}B$ downregulated CD7 gene expression through the p38 MAPK pathway in developing immature thymocytes. However, its regulatory pathway is not well understood in functional mature T-cells. Here, we show that CD7 expression was downregulated by Twist2 in Jurkat cells, a human acute T-cell lymphoma cell line, and in EL4 cells, a mature murine T-cell lymphoma cell line. Furthermore, ectopic expression of Twist2 in Jurkat cells reduced galectin-1-induced apoptosis. While full-length Twist2 decreased CD7 promoter activity, a C-terminal deletion form of Twist2 reversed its inhibition, suggesting an important role of the C-terminus in CD7 regulation. In addition, CD7 expression was enhanced by histone deacetylase inhibitors such as trichostatin A and sodium butyrate, which indicates that Twist2 might be one of candidate factors involved in histone deacetylation. Based on these results, we conclude that upregulation of Twist2 increases the resistance to galectin-1-mediated-apoptosis, which may have significant implications for the progression of some T-cells into tumors such as Sezary cells.

• Journal of Animal Science and Technology
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• v.60 no.5
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• pp.8.1-8.9
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• 2018

Background: Indiscriminate use of antibiotics in livestock and poultry farming has caused emergence of new pathogenic strains. The situation has warrented the development of safe and alternative growth promoters and immunity enhancers in livestock. Herbal additives in animal and bird feed is a centuries-old practice. Thus, the present study investigated the efficacy of a standardized formulation of lipophilic turmeric extract containing curcumin and turmerones, (TF-36), as a natural growth promoter poultry feed additive. Methods: The study was designed on 180 one-day old chicks, assigned into three groups. Control group ($T_0$) kept on basal diet and supplemented groups $T_{0.5}$ and $T_1$ fed with 0.5% and 1% TF-36 fortified basal diet for 42 days. Each dietary group consisted of six replicates of ten birds. Body weight, food intake, food conversion ratio, skin colour, blood biochemical analysis and antioxidant status of serum were investigated. Results: Body weight improved significantly in $T_1$ with a 10% decrease in FCR as compared to the control. TF-36 supplementation in $T_1$ enhanced the antioxidant enzyme activity significantly (p < 0.05) with a decrease (p < 0.05) in lipid peroxidation. It also caused a slight yellow skin pigmentation without any change in meat color, indicating the bioavailability of curcumin from TF-36. However, no significant change in the concentration of serum creatinine, total protein and liver enzyme activities were observed, indicating the safety. Conclusion: In summary, we concluded that TF-36 can be a natural feed additive to improve growth performance in poultry, probably due to the better antioxidant activity and antimicrobial effects contributed by the better bioavailability of curcuminoids and turmerones. Besides, curcuminoids and turmerones were also known to be gastroprotective and anti-inflammatory agents.

• Journal of Life Science
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• v.14 no.1
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• pp.121-125
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• 2004

The synergistic effect of lowered incubation temperature and CroEL/ES expression on the production of soluble form of B. macerans cyclodextrin glucanotransferase (CGTase) was studied in recombinant E. coli. pTCGTl and pGroll carrying the cgt and groEL/ES genes under the control of T7 promoter and pzt-I promoter, respectively, were co-introduced. Tetracycline (10 ng/ml) and IPTG (1 mM) were added at the early-exponential phase (2 hr) and mid-exponential phase (3 hr). Low temperature cultivation at $25^{\circ}C$ with groEL/ES expression improved the activity of CGTase by two fold, compared to $37^{\circ}C$ cultivation without chaperones. SDS-PACE analysis revealed that about 69% of CGTase in the total CGTase protein was found in the soluble fraction by overexpression of GroEL/ES and cultivation at$25^{\circ}C$, whereas 20% of CGTase was detected in the soluble fraction when E. coli was cultivated at $37^{\circ}C$ without chaperone. The amount of soluble CGTase from $25^{\circ}C$ culture with chaperone was 3.5-fold higher than that of $37^{\circ}C$ culture without chaperone. Therefore the expression of CroEL/ES and low temperature cultivation greatly enhanced the soluble production of CGTase in E. coli.

• Journal of Life Science
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• v.12 no.6
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• pp.688-693
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• 2002

Molecular chaperones prevent the misfolding of newly synthesized polypeptides in the cell. The coexpression of molecular chaperones could be expected to improve the production of soluble and active recombinant proteins. In this study, the effect of coexpression of E. coli GroEL/ES chaperone on the active production of Bacillus macerans cyclodextrin glucanotransferase (CGTase) in E. coli was investigated. Two plasmids, pTCGT1 and pGro7 in which the cgt and the groEL/ES genes are under the control of 77 promoter and araB promoter, respectively, were co-transformed into E. coli. With a series of cultures of recombinant E. coli cells, the optimal concentrations of IPTG and L-arabinose were found be 1 mM and 0.3 mg/$m\ell$, respectively. When IPTG and L-arabinose were added at 0.8~1.0 $OD_{600}$ and 0.4~0.5 $OD_{600}$, active CGTase production was increased significantly. This coexpression condition resulted in 1.5-fold increased level of soluble CGTase (0.7~0.73 unit/$m\ell$), compared to the level of CGTase in the single expression (0.36~0.56 unit/$m\ell$). An SDS-PACE analysis revealed that about 33.6% of CGTase in the total CGTase protein was found in the soluble fraction by coexpression of GroEL/ES chaperone.

• International Journal of Industrial Entomology and Biomaterials
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• v.37 no.2
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• pp.49-54
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• 2018

To artificially enhance antimicrobial peptide expression in Bombyx mori, we constructed genetically engineered silkworms overexpressing Rel family transcription factor. The truncated BmRelish1 (BmRelish1t) gene contained a Rel homolog domain (RHD), nuclear localization signal (NLS), acidic and hydrophobic amino acid (AHAA)-rich region, and death domain (DD), but no ankyrin-repeat (ANK) domain. The BmRelish1t gene was controlled by B. mori cytoplasmic actin 3 promoter in the PiggyBac transposon vector. Chromosome analysis of G1 generations of a transgenic silkworm with EGFP expression confirmed stable insertion of BmRelish1t. BmRelish1t gene overexpression in transgenic silkworms resulted in higher mRNA expression levels of B. mori antimicrobial peptides such as lebocin(~20.5-fold), moricin(~8.7-fold), and nuecin(~17.4-fold) than those in normal silkworms.

• Biotechnology and Bioprocess Engineering:BBE
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• v.9 no.5
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• pp.389-392
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• 2004

In order to produce a harmful protein more efficiently, this expression cassette, dubbed pCol-MICT, is directed by the colicin promoter, and was constructed by the insertion of a $rrnBT_1T_2$ fragment of pEXP7, and a MxelnteinCBD fragment of pTXB3, into pSH375. To test whether harmful proteins, including proteolytic enzymes, could be effectively produced by this cassette, the carboxypeptidase (CPase) Taq gene was inserted into the pCol-MICT cassette to yield pCol-CPase Taq-MICT. E coli W3l 10 tells harboring pCol-CPase Taq-MICT produced a large quantity of this enzyme, as much as 47.2 mg of purified from per liter of culture, when cultured in the presence of mitomycin C ($0.4{\mu}g/mL$). This indicates that the colicin promoter-controlled E, coli expression cassette was able to produce almost 8 times of protein than the conventional tar promoter-based system, and that this cassette may be useful in the Synthesis of other harmful proteins.

• Journal of Plant Biotechnology
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• v.34 no.1
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• pp.55-59
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• 2007

The study was carried out to develop transformants resisting to Phyophthora blight disease in the domestic pepper cultivar Subicho. In transforming of syn600 promoter with elicitin gene using Agrobacterium (LBA4404/pBI101 syn600-syn${\alpha}$-elicitin) to cotyledons of pepper, rate of shoot formation in 'Subicho' was 11.1% in medium containing 3 mg/L zeatin and 0.05 mg/L NAA, and also 12.8% in medium containing combination of 4 mg/L zeatin and 0.05 mg/L MAA. For PCR reaction using elicitin gene primer of transformants regenerated from cotyledons, we detected a specific band of 536 bp, and also showed strong signal at position of 536 bp in accordance with NPTII gene used as probe in Southern blot. Transformants pepper shown resistance to blight fungus was inoculated to seedlings of the $T_{1}\;and\;T_{2}$ transformants by concentration (density: zoo spore $10^{3}/mL$).

• Journal of Gastric Cancer
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• v.7 no.1
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• pp.9-15
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• 2007

Purpose: DNA methylation is an important epigenetic factor in tumorigenesis. We hypothesized that polymorphism of the promoter of the DNA methyltransferase 3b (DNMT3b) genes, which are responsible for regulating the methylation status of tumor suppressor genes, are associated with increased risk of gastric cancer. Materials and Methods: In this hospital-based case-control study, to determine the role of this polymorphism of the promoter of DNA methyltransferase 3b (DNMT3b) genes in gastric cancer, we genotyped 176 cases and 70 control subjects. To determine the genotype, we used a polymerase chain reaction restriction fragment length polymorphism assay. We compared alleles and genotypes between the two groups and revealed an association of DNMT3b promoter polymorphism with increased risk of gastric cancer in the Korean population. Results: Genotype frequencies were 14.8% (Cytosine-Cytosine), 71.6% (Cytosine-Thymine), and 13.6% (Thymine- Thymine) in the case patients and 40.0% (Cytosine-Cytosine), 42.9% (Cytosine-Thymine), and 17.1% (Thymine-Thymine) in the control subjects, respectively. Compared with CC homozygotes, CT heterozygotes had a 4.523-fold increased risk (OR, 2.13; 95% CI, 2.324~8.803), and the TT homozygotes had a 2.154-fold elevated risk (OR, 1.42; 95% CI, 0.899~85.165). For the T variant genotype (CT+TT), there was a 3.846-fold increased risk (OR, 1.88; 95% CI, 2.040~7.251). However, no significance was observed in the genotype distributions of both polymorphisms according to histopathology, stage of stomach cancer. The Ssame results were observed with Helicobacter infection. Conclusion: DNMT3b promoter polymorphism, especially the T variant genotype, is associated significantly with thean increased risk of gastric cancer.

• Journal of Microbiology
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• v.34 no.4
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• pp.341-348
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• 1996

The regulation of xylE gene expression was examined by using vector promoter and construction of genetically engineered microorganisms (GEMs) for application in microcosm. When the xylE gene wsa subcloned into pBluscript SK(+) under the control of lac promoter (pTY1) in E. coli, and the expression was induced by IPTG, the enzyme activity of catechol 2, 3-dioxygenase was increased 4.7 times more than that of the crude extracts from transformants harboring pTY1. We suggest that the xylE gene has its own promoter at the upstream portion, because it was able to be expressed even in the absence of IPTG. A recombinant plasmid, pSW3a harboring the xylE gene under the T7 promotor, showed the activity of 14.5 units/mg protein, higher than that of parental strain, E. coli PYT1. The xylE gene in recombinant plasmid pSW3a was used as reporter gene for the application in microcosm ecosystem, since it was used for detection of xylE-positive clones by catechol spray on the agar plates. The pSW3a in E. coli was introduced into Pseudomonas patida to construct GEM strain, and examined for the exxpression and functional stability in microcosms.