• 약학회지
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• 제55권3호
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• pp.203-212
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• 2011

Adipocytes have been known to secrete a number of important proteins called adipokines with roles in energy metabolism, reproduction, cardiovascular function and immunity. In this study we have attempted to identify intensively secretory proteins from 3T3-L1 adipocytes. 3T3-L1 preadipocytes were differentiated into mature adipocytes and then the cells were left in serum-free medium. The supernatant was filtrated and dialyzed. Lyophilized secretome was fractionated by two different methods, 1-D SDS PAGE and RP-FPLC. The tryptic peptides from the gel slices and the FPLC fractions were analyzed by nanoLC/ESI-MS/MS. We identified a total of 303 identical proteins from two methods, 251 proteins from 1-D gel and 184 proteins from RP-FPLC. 86 of them were listed as a secretory protein Finally, we identified many known or unknown secreted proteins existed in the low level including adiponectin, angiotensinogen, bone morphogenetic protein-1 (BMP-1), macrophage migration inhibitory factor (MIF), insulin like growth factor-II (IGF-II), interleukin-6 (IL-6), follistatin-related protein-1, minecan, and resistin. The existence of some of secreted proteins has been confirmed in RNA level. This proteomic experiment is useful for the intensive screening of secretory proteins in many kinds of other cells.

• 한국정보통신학회논문지
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• 제18권10호
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• pp.2562-2570
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• 2014

단백질의 기능을 유추할 수 있는 중요한 정보중의 하나는 단백질이 존재하는 세포내 위치이다. 최근에는 하나의 단백질이 동시에 존재하는 여러 세포내 위치를 예측하는 연구가 활발하다. 본 논문에서는 단백질이 존재하는 세포내의 다중위치를 예측하기 위해서 레이블 멱집합 방법을 개선한다. 레이블 멱집합 방법으로 분류한 다중위치들을 예측 확률에 따라 결합하여 최종적인 다중레이블로 분류한다. 각 다중위치에 대한 정확한 확률적 기여를 구하기 위하여 쌍별 비교와 오류정정 출력코드를 사용한 다중클래스 확률추정 방법을 적용하였다. 단백질 세포내 위치 예측 실험에 제안한 방법을 적용하여 성능이 향상됨을 보였다.

• The Korean Journal of Physiology and Pharmacology
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• 제23권5호
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• pp.411-417
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• 2019

Humanin (HN) is a mitochondrial peptide that exhibits cytoprotective actions against various stresses and diseases. HN has been shown to induce the phosphorylation of AMP-activated protein kinase (AMPK), which is a negative regulator of receptor activator of nuclear factor-${\kappa}B$ ligand (RANKL). However, the role of HN in osteoclastogenesis or other skeletal disorders remains unknown. Here, we examined whether HN regulates osteoclastogenesis via AMPK activation using bone marrow-derived macrophage (BMM) cultures. Our results show that HN inhibited RANKL-induced osteoclast formation and reduced the expression of genes involved in osteoclastogenesis, including nuclear factor of activated T-cells cytoplasmic 1, osteoclastassociated receptor, cathepsin K, and tartrate-resistant acid phosphatase. Moreover, HN increased the levels of phosphorylated AMPK protein; compound C, an AMPK inhibitor, recovered HN-induced osteoclast differentiation. In addition, we found that HN significantly decreased the levels of RANKL-induced reactive oxygen species in BMMs. Therefore, these results indicate that HN plays an important role in osteoclastogenesis and may function as an inhibitor of bone disorders via AMPK activation.

• BMB Reports
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• 제56권9호
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• pp.514-519
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• 2023

Methyltransferase-like 3 (METTL3), a key component of the m⁶A methyltransferase complex, regulates the splicing, nuclear transport, stability, and translation of its target genes. However, the mechanism underlying the regulation of METTL3 expression by alternative splicing (AS) remains unknown. We analyzed the expression pattern of METTL3 after AS in human tissues and confirmed the expression of an isoform retaining introns 8 and 9 (METTL3-IR). We confirmed the different intracellular localizations of METTL3-IR and METTL3 proteins using immunofluorescence microscopy. Furthermore, the endogenous expression of METTL3-IR at the protein level was different from that at the mRNA level. We found that 3'-UTR generation by intron retention (IR) inhibited the export of METTL3-IR mRNA to the cytoplasm, which in turn suppressed protein expression. To the best of our knowledge, this is the first study to confirm the regulation of METTL3 gene expression by AS, providing evidence that the suppression of METTL3 protein expression by IR is an integral part of the mechanism by which 3'-UTR generation regulates protein expression via inhibition of RNA export to the cytoplasm.

• 대한의생명과학회지
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• 제29권4호
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• pp.287-295
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• 2023

Amifostine was developed to protect cells, but it is known to induce cytotoxicity and apoptosis, and the exact mechanism is unknown. In this study, we investigated how the DNA mismatch repair (MMR) system interacts with p53 to prevent apoptosis, cell cycle arrest, and cytoprotective effects induced by amifostine. HCT116 colon cancer cells sublines HCT116/p53+,HCT116/p53+, HCT116/p53-, HCT116/E6 and HCT116+ch3/E6 cells were used for evaluation. Amifostine induced G1 arrest and increased toxicity two-fold in p53- cells regardless of MMR expression. Both G1 cell cycle arrest and induction of p53 protein peaked at 24 h after the start of amifostine exposure. Both G1 cell cycle arrest and induction of p53 protein peaked at 24 h after the start of amifostine exposure. Amifostine induced the expression of p21 protein in both p53+ and p53- cells. As for apoptosis, compared to p53- cells, p53+ cells showed 3.5~4.2 times resistance to amifostine-induced apoptosis. HCT116+E6 with both p53 and MMR loss showed maximum apoptosis at 48 h, and HCT116+ch3/E6HCT116+ch3/E6 with p53 loss showed maximum apoptosis at 24 h. As a result, it was confirmed through in vitro experiments that amifostine-induced G1 cell cycle arrest and apoptosis are mediated through a pathway dependent on MMR and p53 protein.

• Reproductive and Developmental Biology
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• 제35권3호
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• pp.355-361
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• 2011

Ski protein is implicated in proliferation/differentiation in a variety of cells. We had previously reported that Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells, however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to locate Ski protein in the rat ovary during luteinization to predict the possible role of Ski. In order to examine the expression pattern of Ski protein along with the progress of luteinization, follicular growth was induced by administration of equine chorionic gonadotropin to immature female rat, and luteinization was induced by human chorionic gonadotropin treatment to mimic luteinizing hormone (LH) surge. While no Ski-positive granulosa cells were present in preovulatory follicle, Ski protein expression was induced in response to LH surge, and was maintained after the formation of corpus luteum (CL). Though Ski protein is absent in granulosa cells of preovulatory follicle, its mRNA (c-ski) was expressed and the level was unchanged even after LH surge. Taken together, these results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggested that its expression is regulated post-transcriptionally.

• Molecules and Cells
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• 제45권3호
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• pp.148-157
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• 2022

Hepatitis C virus (HCV) is a major cause of chronic liver disease and is highly dependent on cellular proteins for viral propagation. Using protein microarray analysis, we identified 90 cellular proteins as HCV nonstructural 5A (NS5A) interacting partners, and selected telomere length regulation protein (TEN1) for further study. TEN1 forms a heterotrimeric complex with CTC and STN1, which is essential for telomere protection and maintenance. Telomere length decreases in patients with active HCV, chronic liver disease, and hepatocellular carcinoma. However, the molecular mechanism of telomere length shortening in HCV-associated disease is largely unknown. In the present study, protein interactions between NS5A and TEN1 were confirmed by immunoprecipitation assays. Silencing of TEN1 reduced both viral RNA and protein expression levels of HCV, while ectopic expression of the siRNA-resistant TEN1 recovered the viral protein level, suggesting that TEN1 was specifically required for HCV propagation. Importantly, we found that TEN1 is re-localized from the nucleus to the cytoplasm in HCV-infected cells. These data suggest that HCV exploits TEN1 to promote viral propagation and that telomere protection is compromised in HCV-infected cells. Overall, our findings provide mechanistic insight into the telomere shortening in HCV-infected cells.

• 대한약학회:학술대회논문집
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• 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2-2
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• pp.123.2-123.2
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• 2003

The cellular mechanisms by which excess exposure to the excitatory neurotransmitter glutamate can produce neuronal injury are unknown. In this study, we found that glutamate induced cell death at IC (50) of 100 microM on the cultured human SH-SY5Y neuroblastoma cells. It has been hypothesized that glutamate excitotoxicity is related with the elevation of calcium (Ca) levels. To determine the dependence of glutamate neurotoxicity on Ca environment, extracellular (EDTA) and intracellular (BAPTA/AM) chelator were used. (omitted)

• Parasites, Hosts and Diseases
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• 제53권4호
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• pp.403-411
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• 2015

Plasmodium falciparum can invade all stages of red blood cells, while Plasmodium vivax can invade only reticulocytes. Although many P. vivax proteins have been discovered, their functions are largely unknown. Among them, P. vivax reticulocyte binding proteins (PvRBP1 and PvRBP2) recognize and bind to reticulocytes. Both proteins possess a C-terminal hydrophobic transmembrane domain, which drives adhesion to reticulocytes. PvRBP1 and PvRBP2 are large (>326 kDa), which hinders identification of the functional domains. In this study, the complete genome information of the P. vivax RBP family was thoroughly analyzed using a prediction server with bioinformatics data to predict B-cell epitope domains. Eleven pvrbp family genes that included 2 pseudogenes and 9 full or partial length genes were selected and used to express recombinant proteins in a wheat germ cell-free system. The expressed proteins were used to evaluate the humoral immune response with vivax malaria patients and healthy individual serum samples by protein microarray. The recombinant fragments of 9 PvRBP proteins were successfully expressed; the soluble proteins ranged in molecular weight from 16 to 34 kDa. Evaluation of the humoral immune response to each recombinant PvRBP protein indicated a high antigenicity, with 38-88% sensitivity and 100% specificity. Of them, N-terminal parts of PvRBP2c (PVX_090325-1) and PvRBP2 like partial A (PVX_090330-1) elicited high antigenicity. In addition, the PvRBP2-like homologue B (PVX_116930) fragment was newly identified as high antigenicity and may be exploited as a potential antigenic candidate among the PvRBP family. The functional activity of the PvRBP family on merozoite invasion remains unknown.

• Applied Biological Chemistry
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• 제8권
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• pp.11-20
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• 1967

증자대두(蒸煮大豆)에 납두균(納豆菌)(Bacillus subtilis Natto 492-2)을 접종(接種)하고 $38^{\circ}C$에서 발효(醱酵)시키면서 매(每) 10 시간(時間)마다 시료(試料)를 채취(採取) 처리(處理)하여 납두(納豆) 제조과정중(製造過程中)에 일어나는 단백질(蛋白質)의 변화(變化)와 peptide 및 amino acid의 생성양상(生成樣相)에 관(關)하여 다음과 같이 결과(結果)를 얻었다. 1. 증자대두(蒸煮大豆)의 불용성(不溶性) 단백태(蛋白態) 질소(窒素)가 84%에서 발효(醱酵) 50 시간(時間)에 44%로 감소(減少)하였고, 수용성(水溶性) 단백태(蛋白態) 질소(窒素)는 약간(若干)의 증가(增加)를 했지만, peptide, amino acid 및 ammonia 태(態) 질소(窒素)가 8%에서 45%로 증가(增加)하여 단백질(蛋白質)의 가수분해(加水分解)가 현저(顯著)하였다. 2. 납두균(納豆菌)에서 분비(分泌)된 효소(酵素)에 의(依)해 분해(分解)된 peptide 및 amino acid으로 분자사별(分子篩別)한 결과(結果), 대체(大體)로 A.P.L.이 작은 저급(低級) peptide가 많이 생성(生成)됨을 알 수 있으며 발효(醱酵) 30 시간(時間) 이후(以後)에는 이전(以前)보다 단백질(蛋白質) 분해(分解)가 미약(微弱)하게 일어나면서 비교적(比較的) 큰 분자(分子)의 peptide 분해(分解)가 주(主)로 일어나 저급(低級) peptide 및 amino acid 생성(生成)이 많았다. 3. 분자사별(分子篩別)한 X-16 fraction을 시료(試料)로하며 paper chromatography에 의(依)한 amino acid 및 peptide를 동정(同定)한 결과(結果), 16종(種)의 amino acid와 그종(種)의 unknown amino acid가 동정(同定)되었는데, 이들 중(中) phenylalanine은 발효(醱酵) 10 시간(時間) 이후(以後), methionine은 20 시간(時間), proline은 30 시간(時間) 이후(以後)부터 각각(各各) 출현(出現)하였고, 또 Rf치(値)가 다른 3점(點)의 peptide 중(中) 1개(個)는 처음부터 나타나서 발효(醱酵) 20시간이후(時間以後) 소실(消失)하였고, 다른 그점(點)은 20 시간(時間), 40 시간이후(時間以後)부터 각각(各各) 계속(繼續)하여 출현(出現)하였다.