• Korean journal of applied entomology
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• v.36 no.4
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• pp.331-340
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• 1997

Hyalophora cecropia attacin-like antibacterial gene was isolated from Bombyx mori induced with nonpathogenic bacteria. It was expressed in Spodopfera frugiperda 9 (Sf9 cells using baculovirus expression vector system (BEVS), and examined its antibacterial activity. With a cDNA library constructed from fifthinstar B. mori injected with Escherichia coli(4 X IOhcellsllarva), differential screening was performed using naive and induced mRNA probes. BmInc6 clone was screened by partial nucleotide sequence and GenBank database analysis. A complete nucleotide sequence of Bmlnc6 cDNA was determined (GenBank, AF005384). Its insert size was 852 bp and had open reading frame that started translation at position 35 and stopped at 679. And its putative polyadenylational signal existed at 812 bp. The number of amino acid deduced from Bmlnc6 cDNA was 214 and hydropathy analysis showed that this peptide was hydrophilic. This peptide deduced by BmInc6 was named nuecin. When the nuecin gene was expressed in Sf9 cells using BEVS, about 950 bp of the transcripts was detected. In addition, SDS-PAGE analysis showed that the molecular weights of intracellular expressed protein and the mature protein secreted to culture media were approximately 23 and 20 kDa, respectively. The antibacterial activity of nuecin against E. coli and Bacillus subtilis was significantly high, demonstrating that nuecin had a wider antibacterial spectrum with gram negative and positive bacteria than attacin.

• Korean Journal of Ichthyology
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• v.19 no.4
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• pp.266-273
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• 2007

Heat shock proteins (HSPs) are a family of highly conserved proteins playing an important role in the functioning of unstressed and stressed cells. The HSP70 family, the most widely studied of the hsps, is constitutively expressed (hsc70) in unstressed cells and is also induced in response to stressors (hsp70), especially those affecting the protein machinery. The HSP/HSC70 proteins act as molecular chaperones and are crucial for protein functioning, including folding, intracellular localization, regulation, secretion, and protein degradation. Here, we report the identification and characterization of the putative amino acid sequence deduced from one cDNA clone identified as heat shock protein 70. The alignment showed that the putative sequence is 100% identical to the heat shock protein 70 cognate (HSC 70) of olive flounder. The 5'-flanking region sequence (approximately 1 kb) ahead of the hsc70 gene was cloned by genome walking and a putative core promoter region and transcription elements were identified. We characterized the promoter of the olive flounder hsc70 gene by examining the ability of 5'-upstream fragments to drive expression of green fluorescent protein (GFP) in live embryos.

• Korean Journal of Ichthyology
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• v.19 no.4
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• pp.257-265
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• 2007

We constructed a cDNA library of testis from olive flounder (Paralichthys olivaceus) and a total of 248 expressed sequence tag (EST) clones were generated. In order to understand the molecular compositions of the olive flounder testis organs, the expression profiles of the identified clones in the cDNA library were analyzed. Gene annotation procedures and homology searches of the sequenced ESTs were locally done by BLASTX for amino acid similarity comparisons. Of the 248 EST clones, 156 ESTs showed significant homology to previously described genes while 92 ESTs were unidentified or novel. Comparative analysis of the 156 identified ESTs showed that 6 (3.8%) clones were representing 5 unique genes identified as homologous to the previously reported olive flounder ESTs, 100 (64.1%) clones representing 94 unique genes were identified as orthologs of known genes from other organisms, and orthologs were established for 50 (32.1%) clones representing 44 genes of known sequences with unknown functions. Furthermore, the testis library showed a more even distribution of cDNA clones with relatively fewer abundant clones that tend to contribute redundant clones in EST projects; thus, the testis library can supply more unique and novel cDNA sequences in olive flounder EST project.

• Korean Journal of Microbiology
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• v.50 no.2
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• pp.164-168
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• 2014

The ecosystem of the Arctic region has been increasingly affected by global warming. Archaeal ammonia monooxygenase alpha subunit coding gene (amoA) which is a key enzyme for nitrification was used to investigate the effect of runoff water of ice melt on microbial community of nitrogen cycle. The archaeal amoA genes at coastal area of Svalbard, Arctic region were PCR-amplified and sequenced after clone library construction. Analysis of archaeal amoA gene clone libraries suggested that the station 188 which is in the vicinity to the area of runoff water harbor lower ammonia-oxidizing archaeal diversity than the station 176 and 184. The average amino acid sequence identity within all archaeal amoA gene clones was 94% (with 91% nucleotide sequence identity). While all the clones of the station 188 were affiliated with Nitrosoarchaeaum clade containing strains isolated from low-salinity and terrestrial environments, about 45% of total clones of the station 176 and 184 were related to marine Nitosopumilus clade. Interestingly, other typical archaeal amoA gene clones of thaumarchaeal I.1b clade frequently retrieved from terrestrial environments was identified at station 188. Microbial community of nitrogen cycle in marine sediment might be affected by input of sediments caused by runoff glacier melt waters.

• Journal of Genetic Medicine
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• v.16 no.2
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• pp.85-89
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• 2019

Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is a rare autosomal recessive urea cycle disorder. HHH is caused by a deficiency of the mitochondrial ornithine transporter protein, which is encoded by the solute carrier family 25, member 15 (SLC25A15) gene. Recently, government supported Korean newborn screening has been expanded to include a tandem mass spectrometry (MS/MS) measurement of ornithine level. We report a case of a neonate with HHH syndrome showing a normal MS/MS measurement of ornithine level. A female newborn was admitted to neonatal intensive unit due to familial history of HHH syndrome. Her parents were consanguineous Parkistani couple. The subject's older sister was diagnosed with HHH syndrome at age of 30 months based on altered mental status and liver dysfunction. Even though the subject displayed normal ammonia and ornithine levels based on MS/MS analysis, a molecular test confirmed the diagnosis of HHH syndrome. At 1 month of age, amino acid analysis of blood and urine showed high levels of ornithine and homocitrulline. After 11 months of follow up, she showed normal growth and development, whereas affected sister showed progressive cognitive impairment despite no further hyperammonemia after protein restriction and standard therapy. Our report is in agreement with a previous Canadian study, which showed that neonatal samples from HHH syndrome patients demonstrate normal ornithine levels despite having known mutations. Considering the delayed rise of ornithine in affected patients, genetic testing, and repetitive metabolic testing is needed to prevent patient loss in high risk patients.

• Journal of Life Science
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• v.30 no.3
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• pp.304-312
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• 2020

Agarase can be used in the field of basic science, as well as for production of agar-derived high-functional oligosaccharides and bioenergy production using algae. In 2012, we summarized the classification, origin, production, and applications of agar. In this paper, we briefly review the literature on the recombinant expression of agarases from 2012 to the present. Agarase genes originated from 19 genera, including Agarivorans, Flammeovirga, Pseudoalteromonas, Gayadomonas, Catenovulum, Microbulbifer, Cellulophaga, Saccharophagus, Simiduia, and Vibrio. Of the 47 recombinant agarases, there were only two α-agarases, while the rest were β-agarases. All α-agarases produced agarotetraose, while β-agarases yielded many neoagarooligosaccharides ranging from neoagarobiose to neoagarododecaose. The optimum temperature ranged between 25 and 60℃, and the optimum pH ranged from 3.0 to 8.5. There were 14 agarases with an optimum temperature of 50℃ or higher, where agar is in sol state after melting. Artificial mutations, including manipulation of carbohydrate-binding modules (CBM), increased thermostability and simultaneously raised the optimum temperature and activity. Many hosts and secretion signals or riboswitches have been used for recombinant expression. In addition to gene recombination based on the amino acid sequence after agarase purification, recombinant expression of the putative agarase genes after genome sequencing and metagenome-derived agarases have been studied. This study is expected to be actively used in the application fields of agarase and agarase itself.

• Journal of Life Science
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• v.28 no.3
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• pp.351-356
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• 2018

L-Serine dehydratase (LSD) is an iron-sulfur containing enzyme that catalyzes the conversion of L-serine to pyruvate and ammonia. Among the bacterial amino acid dehydratases, it appears that only the L-serine specific enzymes utilize an iron-sulfur cluster at their catalytic site. Moreover, bacterial LSDs are classified into four types based on structural characteristics and domain arrangement. To date, only the LSD enzymes from a few bacterial strains have been studied, but more detailed investigations are required to understand the catalytic mechanism of various bacterial LSDs. In this study, LSD type II from Mycobacterium tuberculosis (MtLSD) H37Rv was expressed and purified to elucidate the biochemical and catalytic properties using the enzyme kinetic method. The L-serine saturation curve of MtLSD exhibited a typically sigmoid character, indicating an allosteric cooperativity. The values of $K_m$ and $k_{cat}$ were estimated to be $59.35{\pm}1.23mM$ and $18.12{\pm}0.20s^{-1}$, respectively. Moreover, the plot of initial velocity versus D-serine concentration at fixed L-serine concentrations showed a non-linear hyperbola decay shape and exhibited a competitive inhibition for D-serine with an apparent $K_i$ value of $30.46{\pm}5.93mM$ and with no change in the $k_{cat}$ value. These results provide insightful biochemical information regarding the catalytic properties and the substrate specificity of MtLSD.

• Journal of Microbiology and Biotechnology
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• v.28 no.1
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• pp.145-156
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• 2018

Most eukaryotic peroxiredoxins (Prxs) are readily inactivated by a high concentration of hydrogen peroxide ($H_2O_2$) during catalysis owing to their "GGLG" and "YF" motifs. However, such oxidative stress sensitive motifs were not found in the previously identified filamentous fungal Prxs. Additionally, the information on filamentous fungal Prxs is limited and fragmentary. Herein, we cloned and gained insight into Aspergillus nidulans Prx (An.PrxA) in the aspects of protein properties, catalysis characteristics, and especially $H_2O_2$ tolerability. Our results indicated that An.PrxA belongs to the newly defined family of typical 2-Cys Prxs with a marked characteristic that the "resolving" cysteine ($C_R$) is invertedly located preceding the "peroxidatic" cysteine ($C_P$) in amino acid sequences. The inverted arrangement of $C_R$ and $C_P$ can only be found among some yeast, bacterial, and filamentous fungal deduced Prxs. The most surprising characteristic of An.PrxA is its extraordinary ability to resist inactivation by extremely high concentrations of $H_2O_2$, even that approaching 600 mM. By screening the $H_2O_2$-inactivation effects on the components of Prx systems, including Trx, Trx reductase (TrxR), and Prx, we ultimately determined that it is the robust filamentous fungal TrxR rather than Trx and Prx that is responsible for the extreme $H_2O_2$ tolerence of the An.PrxA system. This is the first investigation on the effect of the electron donor partner in the $H_2O_2$ tolerability of the Prx system.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.9
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• pp.1245-1252
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• 2017

Objective: Phellodendron amurense (P. amurense) and Humulus japonicus (H. japonicus) are closely involved in anti-oxidative response and increasing antioxidant enzymes activities. However, the effects of their extracts on development of preimplantation bovine embryos have not been investigated. Therefore, we investigated the effects of P. amurense and H. japonicus extracts on developmental competence and quality of preimplantation bovine embryos. Methods: After in vitro fertilization, bovine embryos were cultured for 7 days in Charles Rosenkrans amino acid medium supplemented with P. amurense ($0.01{\mu}g/mL$) and H. japonicus ($0.01{\mu}g/mL$). The effect of this supplementation during in vitro culture on development competence and antioxidant was investigated. Results: We observed that the blastocysts rate was significantly increased (p<0.05) in P. amurense ($28.9%{\pm}2.9%$), H. japonicus ($30.9%{\pm}1.5%$), and a mixture of P. amurense and H. japonicus ($34.8%{\pm}2.1%$) treated groups compared with the control group ($25.4%{\pm}1.6%$). We next confirmed that the intracellular levels of reactive oxygen species (ROS) were significantly decreased (p<0.01) in P. amurense and/or H. japonicus extract treated groups when compared with the control group. Our results also showed that expression of cleaved caspase-3 and apoptotic cells of blastocysts were significantly decreased (p<0.05) in bovine blastocysts derived from both P. amurense and H. japonicus extract treated embryos. Conclusion: These results suggest that proper treatment with P. amurense and H. japonicus extracts in the development of preimplantation bovine embryos improves the quality of blastocysts, which may be related to the reduction of ROS level and apoptosis.

• The Korean Journal of Malacology
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• v.30 no.4
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• pp.399-408
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• 2014

Glutathione S-transferases (GSTs) are a superfamily of detoxification enzymes that primarily catalyze the nucleophilic addition of reduced glutathione to both endogenous and exogenous electrophiles. In this study, we isolated and characterized a full-length of alpha class GST cDNA from the abalone (Haliotis discus hannai). The abalone GST cDNA encodes a 223-amino acid polypeptide with a calculated molecular mass of 25.8 kDa and isoelectric point of 5.69. Multiple alignments and phylogenetic analysis with the deduced abalone GST protein revealed that it belongs to the alpha class GSTs and showed strong homology with disk abalone (Haliotis discus discus) putative alpha class GST. Abalone GST mRNA was ubiquitously detected in all tested tissues. GST mRNA expression was comparatively high in the mantle, gill, liver, and digestive duct, however, lowest in the hemocytes. Expression level of abalone GST mRNA in the mantle, gill, liver, and digestive duct was 182.7-fold, 114.8-fold, 4675.8-fold, 406.1-fold higher than in the hemocytes, respectively. Expression level of abalone GST mRNA in the liver was peaked at 6 h post-infection with Vibrio parahemolyticus and decreased at 12 h post-infection. While the expression level of abalone GST mRNA in the hemocytes was drastically increased at 3 h post-infection with Vibrio parahemolyticus. These results suggest that abalone GST is conserved through evolution and may play roles similar to its mammalian counterparts.