• Journal of Applied Biological Chemistry
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• v.49 no.1
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• pp.1-7
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• 2006

Differential display reverse transcription PCR (DDRT-PCR) analysis was performed on lily 'Marcopolo' bulb scale for isolation of expressed genes during bulblet formation. Cu/Zn lily-superoxide dismutase (LSOD) of 872 bp gene, with ability to scavenge reactive oxygen in stress environment, was isolated. Northern blot analysis showed expression levels of LSOD maximized 12 days after bulblet formation. Ti plasmid vectors were constructed with sense and antisense expressions of LSOD gene and transformed into potato. Southern blot analysis of transgenic potatoes revealed different copies of T-DNA were incorporated into potato genome. In transgenic potatoes, lily SOD gene was overexpressed in sense lines and not in antisense lines. In native polyacrylamide gel electrophoresis analysis, additional engineered LSOD was detected in sense overexpressed transgenic line only. Transgenic potatoes were subjected to oxidative stress, such as herbicide methyl viologen (MV). Transgenic potato lines with sense orientation exhibited increased tolerance to MV, whereas in antisense lines exhibited decreased tolerance. In vitro tuberization of transgenic potato with sense orientation was promoted, but was inhibited in transgenic potato with antisense orientation.

• BMB Reports
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• v.52 no.11
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• pp.665-670
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• 2019

Nucleotide-binding oligomerization domain protein 2 (NOD2), an intracellular pattern recognition receptor, plays important roles in inflammation and cell death. Previously, we have shown that NOD2 is expressed in vascular smooth muscle cells (VSMCs) and that NOD2 deficiency promotes VSMC proliferation, migration, and neointimal formation after vascular injury. However, its role in endoplasmic reticulum (ER) stress-induced cell death in VSMCs remains unclear. Thus, the objective of this study was to evaluate ER stress-induced viability of mouse primary VSMCs. NOD2 deficiency increased ER stress-induced cell death and expression levels of apoptosis mediators (cleaved caspase-3, Bax, and Bak) in VSMCs in the presence of tunicamycin (TM), an ER stress inducer. In contrast, ER stress-induced cell death and expression levels of apoptosis mediators (cleaved caspase-3, Bax, and Bak) were decreased in NOD2-overexpressed VSMCs. We found that the $IRE-1{\alpha}-XBP1$ pathway, one of unfolded protein response branches, was decreased in NOD2-deficient VSMCs and reversed in NOD2-overexpressed VSMCs in the presence of TM. Furthermore, NOD2 deficiency reduced the expression of XBP1 target genes such as GRP78, PDI-1, and Herpud1, thus improving cell survival. Taken together, these data suggest that the induction of ER stress through NOD2 expression can protect against TM-induced cell death in VSMCs. These results may contribute to a new paradigm in vascular homeostasis.

• Journal of Microbiology and Biotechnology
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• v.31 no.11
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• pp.1545-1551
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• 2021

Exopolysaccharides (EPSs) such as capsular polysaccharide (CPS) are important bioactive carbohydrate compounds and are often used as bioenrichment agents and bioabsorbers to remove environmental pollutants like di-n-butyl phthalate (DBP). Among the EPS-producing bacteria, lactic acid bacteria (LAB) have gained the most attention. As generally recognized as safe (GRAS) microorganisms, LAB can produce EPSs having many different structures and no health risks. However, EPS production by LAB does not meet the needs of large-scale application on an industrial scale. Here, the capA gene (encoding CPS biosynthesis protein) was overexpressed in Lactobacillus plantarum P1 to improve the production of EPSs and further enhance the DBP adsorption capability. Compared with P1, the CPS production in capA overexpressed strain was increased by 11.3 mg/l, and the EPS thickness was increased from 0.0786 ± 0.0224 ㎛ in P1 to 0.1160 ± 0.0480 ㎛ in P1-capA. These increases caused the DBP adsorption ratio of P1-capA to be doubled. Overall, the findings in this study provide a safe method for the adsorption and removal of DBP.

• Proceedings of the Korean Society of Crop Science Conference
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• 2022.10a
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• pp.133-133
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• 2022

Temperature is a crucial environmental factor for rice cultivation due to the climate change and can influence the rice growth and development. Therefore, the effect of temperature on plant growth characters was examined during the vegetative stage. Plants were grown under three different temperatures: 23℃/13℃ for 18℃, 26℃/16℃ for 21℃ and 29℃/19℃ for 24℃ in the phytotron. The temperature was treated after transplanting and ended in early panicle initiation stage. Heading date of the two varieties were strongly affected by the temperature and were delayed in the 18℃. The plant height in the 18℃ was 21 % shorter than the 21℃ and 24℃ and the tiller and leaf number were increased in the 18℃. All the growth rates of the characters were the slowest in 18℃. The stem dry weight was significantly increased in 18℃. Nitrogen content was increased in the leaves of 18℃ whereas available phosphate and potassium content was found to be increased in the stems of 21℃ and 24℃. OsNRT 2.1 was overexpressed in the leaves and stems of 18℃ and OsNRT2.3a could be expressed in 18℃ and 21℃ temperatures whereas more expressed in 21℃. OsPT1 and OsPT6 could be expressed in the leaf of 18℃ and 24℃ but could be expressed in the stem of 18℃. OsHAK1 and OsHAK5 could be overexpressed in the leaves and stems of 18℃. For hormone, OsCKX2 gene was found to be up regulated in the leaves of 18℃ and OsIAA1 gene could be expressed more in the stem of 24℃.

• Molecules and Cells
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• v.46 no.12
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• pp.764-777
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• 2023

Recombinant immunotoxins (RITs) are fusion proteins consisting of a targeting domain linked to a toxin, offering a highly specific therapeutic strategy for cancer treatment. In this study, we engineered and characterized RITs aimed at mesothelin, a cell surface glycoprotein overexpressed in various malignancies. Through an extensive screening of a large nanobody library, four mesothelin-specific nanobodies were selected and genetically fused to a truncated Pseudomonas exotoxin (PE24B). Various optimizations, including the incorporation of furin cleavage sites, maltose-binding protein tags, and tobacco etch virus protease cleavage sites, were implemented to improve protein expression, solubility, and purification. The RITs were successfully overexpressed in Escherichia coli, achieving high solubility and purity post-purification. In vitro cytotoxicity assays on gastric carcinoma cell lines NCI-N87 and AGS revealed that Meso(Nb2)-PE24B demonstrated the highest cytotoxic efficacy, warranting further characterization. This RIT also displayed selective binding to human and monkey mesothelins but not to mouse mesothelin. The competitive binding assays between different RIT constructs revealed significant alterations in IC₅₀ values, emphasizing the importance of nanobody specificity. Finally, a modification in the endoplasmic reticulum retention signal at the C-terminus further augmented its cytotoxic activity. Our findings offer valuable insights into the design and optimization of RITs, showcasing the potential of Meso(Nb2)-PE24B as a promising therapeutic candidate for targeted cancer treatment.

• Fisheries and Aquatic Sciences
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• v.2 no.1
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• pp.93-97
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• 1999

The promoter region of alginate lyase gene (aly) from Pseudomonas sp. W7 was used for the high expression of gilthead seabream (Sparus aurata) growth hormone (GH) gene in Esherichia coli. PCR product encoding the premature segment of the growth hormone. was cloned to the downstream of aly promoter. GH was overexpressed With 46 ammo acid of alginate lyase as fusion protein. GH was immunoreactive and production of GH was repressed with supplementation of $0.4\%$ glucose into culture media.

• Journal of Microbiology and Biotechnology
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• v.17 no.12
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• pp.2066-2070
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• 2007

It was found that Shinorhizobium meliloti hemoprotein (SM) was more effective than Vitreoscilla hemoglobin (Vhb) in promoting secondary metabolites production when overexpressed in Streptomyces lividans TK24. The transformant with sm (sm-transformant) produced 2.7-times and 3-times larger amounts of actinorhodin than the vhb-transformant in solid culture and flask culture, respectively. In both solid and flask cultures, a larger amount of undecylprodigiocin was produced by the sm-transformant. It is considered that the overexpression of SM especially has activated the pentose phosphate pathway through oxidative stress, as evidenced by an increased NADPH production observed, and that it has promoted secondary metabolites biosynthesis.

• 한국생물공학회:학술대회논문집
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• 2001.11a
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• pp.367-369
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• 2001

Recombinant lipase gene (pYEGA ${\alpha}$ -lip) originated from Bacillus stearothermophillus Ll was overexpressed in Saccharomyces cerevisiae. The lipase gene expression level was compared by controlling a constant specifjc growth rates( ${\mu}$ = 0.03, 0.05, 0.07 and $0.1h^{-1}$. Cell g개wth was successfully controlled at the desired rates by feeding rate of glucose and the formation of by-product or accumulation of the glucose was not observed. Above the growth rate of $0.1h^{-1}$. the desired growth rate could not be achieved caused accumulation of by-products(ethanol). The lipase production increased as the specific growth rate decreased. The specific production rate at the lowest specific growth rater(${\mu}$ =0.03) was above 2- folds than the others.

• BMB Reports
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• v.32 no.2
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• pp.147-153
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• 1999

Previously, we reported that the prothrombin kringle 2 (fragment 2), induced by LPS administration into rabbit, inhibited bFGF-stimulated BCE cell growth (Lee et al., 1998). In this study, we cloned and overexpressed the kringle 2 domain of rabbit and human prothrombin as a fusion protein with the pelB leader sequence in E. coli using the T7 promoter. The fusion protein was cleaved during translocation into the peri plasmic space, and cleaved recombinant protein was readily isolated from whole cell lysate by DEAE-Sepharose and Sephacryl S-200 gel filtration chromatography. Both the recombinant rabbit and human prothrombin kringle 2 showed very similar biochemical and functional characteristics to the rabbit prothrombin kringle 2 purified from rabbit serum, in terms of abnormal electrophoretic migration and endothelial cell growth inhibitory activity.

• BMB Reports
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• v.44 no.10
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• pp.692-697
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• 2011

To investigate the pathways of oxidoreductases in plants, 2 key components in thioredox systems i.e. thioredoxin h (Trx h) and NADPH-dependent thioredoxin reductase (NTR) genes were first isolated from tomatoes (Solanum lycopersicum). Subsequently, the coding sequences of Trx h and NTR were inserted into pET expression vectors, and overexpressed in Escherichia coli. In the UV-Visible spectra of the purified proteins, tomato Trx h was shown to have a characteristic 'shoulder' at ~290 nm, while the NTR protein had the 3 typical peaks unique to flavoenzymes. The activities of both proteins were demonstrated by following insulin reduction, as well as DTNB reduction. Moreover, both NADPH and NADH could serve as substrates in the NTR reduction system, but the catalytic efficiency of NTR with NADPH was 2500-fold higher than with NADH. Additionally, our results reveal that the tomato Trx system might be involved in oxidative stress, but not in cold damage.