• Bulletin of the Korean Chemical Society
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• 제31권6호
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• pp.1519-1526
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• 2010

The behavior of peptide or protein solutes in saline aqueous solution is a fundamental topic in physical chemistry. Addition of ions can strongly alter the thermodynamic and physical properties of peptide molecules in solution. In order to study the effects of added ionic salts on protein conformation and dynamics, we have used the molecular dynamics (MD) simulations to investigate the behavior of Staphylococcus aureus Hfq protein under two different ionic concentrations: 0.1 M NaCl and 1.0 M NaCl in presence and absence of RNA (a hepta-oligoribonucleotide AU5G). Hfq, a global regulator of gene expression is highly conserved and abundant RNA-binding protein. It is already reported that in vivo the increase of ionic strength results in a drastic reduction of Hfq affinity for $Q{\beta}$ RNA and reduces the tendency of aggregation of Escherichia coli host factor hexamers. Our results revealed the crucial role of 0.1 M NaCl Hfq system on the bases with strong hydrogen bonding interactions and by stabilizing the aromatic stacking of Tyr42 residue of the adjacent subunits/monomers with the adenine and uridine nucleobases. An increase in RNA pore diameter and weakened compactness of the Hfq-RNA complex was clearly observed in 1.0 M NaCl Hfq system with bound RNA. Aggregation of monomers in Hfq and the interaction of Hfq with RNA are greatly affected due to the presence of high ionic strength. Higher the ionic concentration, weaker is the aggregation and interaction. Our results were compatible with the experimental data and this is the first theoretical report for the experimental study done in 1980 by Uhlenbeck group for the present system.

• BMB Reports
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• 제56권11호
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• pp.600-605
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• 2023

Intrahepatic cholangiocarcinoma (ICC) is a bile duct cancer and a rare malignant tumor with a poor prognosis owing to the lack of an early diagnosis and resistance to conventional chemotherapy. A combination of gemcitabine and cisplatin is the typically attempted first-line treatment approach. However, the underlying mechanism of resistance to chemotherapy is poorly understood. We addressed this by studying dynamics in the human ICC SCK cell line. Here, we report that the regulation of glucose and glutamine metabolism was a key factor in overcoming cisplatin resistance in SCK cells. RNA sequencing analysis revealed a high enrichment cell cycle-related gene set score in cisplatin-resistant SCK (SCK-R) cells compared to parental SCK (SCK WT) cells. Cell cycle progression correlates with increased nutrient requirement and cancer proliferation or metastasis. Commonly, cancer cells are dependent upon glucose and glutamine availability for survival and proliferation. Indeed, we observed the increased expression of GLUT (glucose transporter), ASCT2 (glutamine transporter), and cancer progression markers in SCK-R cells. Thus, we inhibited enhanced metabolic reprogramming in SCK-R cells through nutrient starvation. SCK-R cells were sensitized to cisplatin, especially under glucose starvation. Glutaminase-1 (GLS1), which is a mitochondrial enzyme involved in tumorigenesis and progression in cancer cells, was upregulated in SCK-R cells. Targeting GLS1 with the GLS1 inhibitor CB-839 (telaglenastat) effectively reduced the expression of cancer progression markers. Taken together, our study results suggest that a combination of GLUT inhibition, which mimics glucose starvation, and GLS1 inhibition could be a therapeutic strategy to increase the chemosensitivity of ICC.

• 대한의생명과학회지
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• 제13권4호
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• pp.251-261
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• 2007

A myofiber of skeletal muscle is composed of myofibrils, sarcolemma (plasma membrane), and constameres, which anchor the myofibrils to the sarcolemma. Achvillin is a recently identified F-actin binding muscle protein, co-isolates with dystrophin and caveolin-3 in low-density sarcolemma of striated muscle, and colocalizes with dystrophin at costameres, the specialized adhesion sites in muscle. Archvillin also binds to nebulin and localizes at myofibrillar Z-discs, the lateral boundaries of the sarcomere in muscle. However other roles of archvillin on the dynamics of myofibrillogenesis remain to be defined. The goal of this study is, by using siRNA-mediated gene silencing technique, to investigate the effect of archvillin on the dynamics of myofibrillogenesis in cell culture of a mouse skeletal myogenic cell line (C2C12), where presumptive myoblasts withdraw from the cell cycle, fuse, undergo de novo myofibrillogenesis, and differentiate into mature myotubes. The roles of archvillin in the assembly and maintenance of myofibril and during the progression of myofibrillogenesis induced in skeletal myoblast following gene silencing in the cell culture were investigated. Fluorescence microscopy demonstrated that the distribution of archvillin was changed along the course of myofibril assembly with nebulin, vinculin and F-actin and then located at Z-lines with nebulin. Fluorescence microscopy demonstrated that knockdown of mouse archvillin expression led to an impaired assembly of new myofibrillar clusters and delayed fusion and myofibrillogenesis although the mouse archvillin siRNA did not affect those expressions of archvillin binding proteins, such as nebulin and F-actin. This result is corresponded with that of RT-PCR and western blots. When the perturbed archvillin was rescued by co-transfection with GFP or Red tagged human archvillin construct, the inhibited cell fusion and myotube formation was recovered. By using siRNA technique, archvillin was found to be involved in early stage of myofibrillogenesis. Therefore, the current data suggest the idea that archvillin plays critical roles on cell fusion and dynamic myofibril assembly.

• 생명과학회지
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• 제33권11호
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• pp.851-858
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• 2023

Unique cartilage matrix-associated protein (UCMA)은 γ-카르복실화(Gla) 잔기가 풍부한 간외 비타민 K 의존 단백질이다. UCMA는 조골세포 분화를 촉진하고 뼈 형성을 강화한다고 보고되고 있지만 고혈당 스트레스 하에서 조골세포에 미치는 영향에 대해서는 아직 알려진 바가 없다. 본 연구에서는 고혈당 조건하에서의 MC3T3-E1 조골세포에서 UCMA 효과를 조사하기 위해 MC3T3-E1 조골세포를 높은 포도당에 노출한 후 재조합 UCMA 단백질을 처리하였다. MC3T3-E1 세포에서 활성 산소종(ROS)의 생성은 고혈당 조건하에서 증가했으나 UCMA 단백질 처리 후 감소했음을 CellROX 및 MitoSOX 염색으로 확인하였다. 또한 고혈당 조건에서 UCMA 단백질을 함께 처리한 MC3T3-E1 세포에서 정량적 중합효소 연쇄반응 결과, 항산화 유전자인 nuclear factor erythroid 2-related factor 2 와 superoxide dismutase 1 발현이 증가하였다. 동일 조건하에서 UCMA 단백질 처리에 의해 heme oxygenase-1 발현 감소와 함께 세포질에서 핵으로의 전위가 감소되었고, 미토콘드리아 분열에 관여하는 dynamin-related protein 1 발현이 증가하였으며, AKT 신호 활성은 억제되었다. 종합적으로 UCMA는 고혈당에 노출된 조골세포에서 ROS 생성을 완화하고, 항산화 유전자 발현을 증가시키고, 미토콘드리아 역학에 영향을 미치며, AKT 신호를 조절하는 것으로 보인다. 본 연구는 UCMA의 세포 메커니즘에 대한 이해를 돕고, 대사 장애 관련한 골 합병증에 대한 새로운 치료제로서의 잠재적 사용 가능성을 제시하고 있다.

• 한국지능시스템학회논문지
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• 제25권6호
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• pp.529-535
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• 2015

지금까지 생체 네트워크 분석 연구는 정적(static)인 개념으로만 다루어졌다. 그러나 실제 생명현상이 발생하는 세포 내에서는 세포의 상태 및 외부 환경에 따라 일부 단백질과 그 상호작용만이 선택적으로 활성화된다. 따라서 생체 네트워크의 구조가 시간의 흐름에 따라 변화하는 동적(dynamic)인 개념이 적용되어야 하며, 이런 개념은 질병의 진행 추이를 분석하는데 효율적이다. 본 논문에서는 동적인 네트워크 방법을 알츠하이머 질병에 적용하여 질병이 진행되는 단계에 따라 변화하는 단백질 상호작용 네트워크의 구조적, 기능적 특징에 대하여 분석하고자 한다. 우선, 유전자 발현데이터를 기반으로 각 질병의 진행 상태에 따른 부분 네트워크(정상, 초기, 중기, 말기)를 구축하였다. 이를 기반으로, 네트워크의 구조적 특성 분석을 수행하였다. 또한 기능적 특성 분석을 위해 유전자 군집(module)을 탐색하고, 군집별 유전자 기능(Gene Ontology) 분석을 수행했다. 그 결과, 네트워크의 특성들은 각 질병의 단계와 잘 대응되며, 동적 네트워크 분석법이 중요한 생물학적 이벤트를 설명하는데 이용될 수 있음을 보였다. 결론적으로 제안된 연구 방법을 통하여 그동안 알려지지 않았던 질병유발에 관련된 주요 네트워크 변화를 관측할 수 있고, 질병에 관여하는 복잡한 분자 수준의 발생 기작과 진행 과정을 이해하는데 중요한 정보를 획득할 수 있다.

• BMB Reports
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• 제40권5호
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• pp.723-730
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• 2007

Kinesin is an ATP-driven microtubule motor protein that plays important roles in control of microtubule dynamics, intracellular transport, cell division and signal transduction. The kinesin superfamily is composed of numerous members that are classified into 14 subfamilies. Animal kinesins have been well characterized. In contrast, plant kinesins have not yet to be characterized adequately. Here, a novel plant-specific kinesin gene, GhKCH2, has been cloned from cotton (Gossypium hirsutum) fibers and biochemically identified by prokaryotic expression, affinity purification, ATPase activity assay and microtubule-binding analysis. The putative motor domain of GhKCH2, $M_{396-734}$ corresponding to amino acids Q396-N734 was fused with 6$\times$His-tag, soluble-expressed in E. coli and affinity-purified in a large amount. The biochemical analysis demonstrated that the basal ATPase activity of $M_{396-734}$ is not activated by $Ca^{2+}$, but stimulated 30-fold max by microtubules. The enzymatic activation is microtubule-concentration-dependent, and the concentration of microtubules that corresponds to half-maximum activation was about 11 ${\mu}M$, much higher than that of other kinesins reported. The cosedimentation assay indicated that $M_{396-734}$ could bind to microtubules in vitro whenever the nucleotide AMP-PNP is present or absent. As a plant-specific microtubule-dependent kinesin with a lower microtubule-affinity and a nucleotide-independent microtubule-binding ability, cotton GhKCH2 might be involved in the function of microtubules during the deposition of cellulose microfibrils in fibers or the formation of cell wall.

• Asian-Australasian Journal of Animal Sciences
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• 제12권1호
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• pp.129-138
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• 1999

If rumen bacteria can be manipulated to utilize nutrients (i.e., ammonia and plant cell wall carbohydrates) more completely and efficiently, the need for protein supplementation can be reduced or eliminated and the digestion of fiber in forage or agricultural residue-based diets could be enhanced. However, these approaches require a complete and accurate description of the rumen community, as well as methods for the rapid and accurate detection of microbial density, diversity, phylogeny, and gene expression. Molecular ecology techniques based on small subunit (SSU) rRNA sequences, nucleic acid probes and the polymerase chain reaction (PCR) can potentially provide a complete description of the microbial ecology of the rumen of ruminant animals. The development of these molecular tools will result in greater insights into community structure and activity of gut microbial ecosystems in relation to functional interactions between different bacteria, spatial and temporal relationships between different microorganisms and between microorganisms and reed panicles. Molecular approaches based on SSU rRNA serve to evaluate the presence of specific sequences in the community and provide a link between knowledge obtained from pure cultures and the microbial populations they represent in the rumen. The successful development and application of these methods promises to provide opportunities to link distribution and identity of gastrointestinal microbes in their natural environment with their genetic potential and in situ activities. The use of approaches for assessing pupulation dynamics as well as for assessing community functionality will result in an increased understanding and a complete description of the gastrointestinal communities of production animals fed under different dietary regimes, and lead to new strategies for improving animal growth.

• IMMUNE NETWORK
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• 제14권2호
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• pp.89-99
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• 2014

Graft-versus-host disease (GVHD) is a fatal complication that occurs after allogeneic hematopoietic stem cell transplantation. To understand the dynamics of CD4 and CD8 T cell production of IFN-${\gamma}$ and IL-17 during GVHD progression, we established a GVHD model by transplanting T cell-depleted bone marrow (TCD-BM) and purified T cells from B6 mice into irradiated BALB.B, creating an MHC-matched but minor histocompatibility (H) antigen-mismatched transplantation (B6 ${\rightarrow}$ BALB.B GVHD). Transplantation-induced GVHD was confirmed by the presence of the appropriate compositional changes in the T cell compartments and innate immune cells in the blood and the systemic secretion of inflammatory cytokines. Using this B6 ${\rightarrow}$ BALB.B GVHD model, we showed that the production of IFN-${\gamma}$ and IL-17 by CD4 T cells preceded that by CD8 T cells in the spleen, mesenteric lymph node, liver, and lung in the BALB.B GVHD host, and Th1 differentiation predated Th17 differentiation in all organs during GVHD progression. Such changes in cytokine production were based on changes in cytokine gene expression by the T cells at different time points during GVHD development. These results demonstrate that both IFN-${\gamma}$ and IL-17 are produced by CD4 and CD8 T cells but with different kinetics during GVHD progression.

• 전자공학회논문지S
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• 제36S권12호
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• pp.10-19
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• 1999

셀룰라 오토마타 신경망(CANS)은 생물학적 발생과 진화에 기반한 신경망 모델이다. CANS에서 각 뉴런은 상호간에 국소적인 연결을 갖고 있으며 카오스 뉴런 모델의 동작 방정식에 따라 펄스의 형태로 동작한다. 신경망은 초기 패턴을 셀룰라 오토마타(CA) 규칙에 따라 발생시켜 얻어진다. 기존의 연구에서는 유용한 기능을 얻기 위하여 초기패턴을 진화시켰다. 그러나 이 방법은 신경망의 표현공간을 모두 나타낼 수 없다. 따라서 본 논문에서는 신경망의 표현공간이 작아지는 문제점을 개선하기 위한 CA의 발생규칙을 진화시키는 방법을 제안한다. DNA 코딩은 코딩의 중복과 여분을 효과적으로 사용하며 규칙의 표현에 매우 적합하다. 본 논문에서는 CA 규칙의 일반적인 표현방법을 제시하고 DNA 코드를 CA 규칙으로 해석하는 방법을 제안한다. 제안된 방법은 자율이동로봇의 제어기에 사용하여 주행 문제에 적용함으로써 그 유효성을 확인하였다.

• Mass Spectrometry Letters
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• 제3권4호
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• pp.93-100
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• 2012

Cellular processes such as proliferation, differentiation, and adaptation to environmental changes are regulated by protein phosphorylation. In order to enhance the understanding of molecular dynamics for biological process in detail, it is necessary to develop sensitive and comprehensive analytical methods for the determination of protein phosphorylation. Neural stem cells hold great promise for neural repair following an injury or disease. In this study, we made differentiated oligodendrocytes from human neural stem cells using over-expression of olig2 gene. We confirmed using quantitative phosphoproteome analysis approach that combines stable isotope labeling by amino acids in cell culture (SILAC) and $TiO_2$ micro-column for phosphopeptide enrichment with $MS^2$ and $MS^3$ mass spectrometry. We detected 275 phosphopeptides which were modulated at least 2-fold between human neural stem cells and oligodendrocytes. Among them, 23 phosphoproteins were up-regulated in oligodendrocytes and 79 phosphoproteins were up-regulated in F3 cells.