• Title/Summary/Keyword: Target Fusion

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Identification and Characterization of Protein Arginine Methyltransferase 1 in Acanthamoeba castellanii

  • Moon, Eun-Kyung;Kong, Hyun-Hee;Hong, Yeonchul;Lee, Hae-Ahm;Quan, Fu-Shi
    • Parasites, Hosts and Diseases
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    • v.55 no.2
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    • pp.109-114
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    • 2017
  • Protein arginine methyltransferase (PRMT) is an important epigenetic regulator in eukaryotic cells. During encystation, an essential process for Acanthamoeba survival, the expression of a lot of genes involved in the encystation process has to be regulated in order to be induced or inhibited. However, the regulation mechanism of these genes is yet unknown. In this study, the full-length 1,059 bp cDNA sequence of Acanthamoeba castellanii PRMT1 (AcPRMT1) was cloned for the first time. The AcPRMT1 protein comprised of 352 amino acids with a SAM-dependent methyltransferase PRMT-type domain. The expression level of AcPRMT1 was highly increased during encystation of A. castellanii. The EGFP-AcPRMT1 fusion protein was distributed over the cytoplasm, but it was mainly localized in the nucleus of Acanthamoeba. Knock down of AcPRMT1 by synthetic siRNA with a complementary sequence failed to form mature cysts. These findings suggested that AcPRMT1 plays a critical role in the regulation of encystation of A. castellanii. The target gene of AcPRMT1 regulation and the detailed mechanisms need to be investigated by further studies.

In Vitro Selection of High Affinity DNA-Binding Protein Based on Plasmid Display Technology

  • Choi, Yoo-Seong;Joo, Hyun;Yoo, Young-Je
    • Journal of Microbiology and Biotechnology
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    • v.15 no.5
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    • pp.1022-1027
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    • 2005
  • Based on plasmid display technology by the complexes of fusion protein and the encoding plasmid DNA, an in vitro selection method for high affinity DNA-binding protein was developed and experimentally demonstrated. The GAL4 DNA-binding domain (GAL4 DBD) was selected as a model DNA-binding protein, and enhanced green fluorescent protein (EGFP) was used as an expression reporter for the selection of target proteins. Error prone PCR was conducted to construct a mutant library of the model. Based on the affinity decrease with increased salt concentration, mutants of GAL4 DBD having high affinity were selected from the mutant protein library of protein-encoding plasmid complex by this method. Two mutants of (Lys33Glu, Arg123Lys, Ile127Lys) and (Ser47Pro, Ser85Pro) having high affinity were obtained from the first generation mutants. This method can be used for rapid in vitro selection of high affinity DNA-binding proteins, and has high potential for the screening of high affinity DNA-binding proteins in a sequence-specific manner.

Characterization of HEK293 and Namalwa Cell Cultures by Using Design of Experiment (실험계획법을 이용한 HEK293 및 Namalwa 세포배양 특성 규명)

  • Kang, Kyung-Ho;Seo, Joon-Serk;Kim, Dong-Il
    • KSBB Journal
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    • v.27 no.3
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    • pp.186-194
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    • 2012
  • Various human host cell lines, which are more effective than the other original human cell lines, have been developed and used. Highly efficient human cell line can be obtained from the fusion between human embryonic kidney 293 (HEK293) and human Burkitt's lymphoma cells (Namalwa). Fused cell line has the advantages of both cell lines such as the high transfection efficacy of HEK293 cells and the constitutive expression of Epstein-Barr virus (EBV) genome which is related with high expression of target protein and anti-apoptotic growth of Namalwa cells. In this study, characterization of two original cell lines was performed by using design of experiment (DOE) considering cell maintenance, media development, optimization of culture condition, and scale-up. The formation of aggregates was apparent with high glutamine concentration at more than 6 mM. Supplementation of hydrolysates showed positive effects on the growth performances of HEK293 cells. On the contrary, Namalwa cells showed negative results. It was confirmed that Namalwa cells were more sensitive to lower temperature at $35^{\circ}C$ and hyperosmotic condition over 260 mOsm/kg. In addition, both cell lines showed limited growth in 3-L bioreactor due to shear stress.

Chronic Myeloid Leukemia - Prognostic Value of Mutations

  • Kaleem, Bushra;Shahab, Sadaf;Ahmed, Nuzhat;Shamsi, Tahir Sultan
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.17
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    • pp.7415-7423
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    • 2015
  • Chronic myeloid leukemia (CML) is a stem cell disorder characterized by unrestricted proliferation of the myeloid series that occurs due to the BCR-ABL fusion oncogene as a result of reciprocal translocation t(9;22) (q34;q11). This discovery has made this particular domain a target for future efforts to cure CML. Imatinib revolutionized the treatment options for CML and gave encouraging results both in case of safety as well as tolerability profile as compared to agents such as hydroxyurea or busulfan given before Imatinib. However, about 2-4% of patients show resistance and mutations have been found to be one of the reasons for its development. European Leukemianet gives recommendations for BCR-ABL mutational analysis along with other tyrosine kinase inhibitors (TKIs) that should be administered according to the mutations harbored in a patient. The following overview gives recommendations for monitoring patients on the basis of their mutational status.

A Study on Emotion Recognition Systems based on the Probabilistic Relational Model Between Facial Expressions and Physiological Responses (생리적 내재반응 및 얼굴표정 간 확률 관계 모델 기반의 감정인식 시스템에 관한 연구)

  • Ko, Kwang-Eun;Sim, Kwee-Bo
    • Journal of Institute of Control, Robotics and Systems
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    • v.19 no.6
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    • pp.513-519
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    • 2013
  • The current vision-based approaches for emotion recognition, such as facial expression analysis, have many technical limitations in real circumstances, and are not suitable for applications that use them solely in practical environments. In this paper, we propose an approach for emotion recognition by combining extrinsic representations and intrinsic activities among the natural responses of humans which are given specific imuli for inducing emotional states. The intrinsic activities can be used to compensate the uncertainty of extrinsic representations of emotional states. This combination is done by using PRMs (Probabilistic Relational Models) which are extent version of bayesian networks and are learned by greedy-search algorithms and expectation-maximization algorithms. Previous research of facial expression-related extrinsic emotion features and physiological signal-based intrinsic emotion features are combined into the attributes of the PRMs in the emotion recognition domain. The maximum likelihood estimation with the given dependency structure and estimated parameter set is used to classify the label of the target emotional states.

Tyrosine Kinase Inhibitors in Ph+ Chronic Myeloid Leukemia Therapy: a Review

  • Shah, Krupa;Parikh, Sonia;Rawal, Rakesh
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.7
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    • pp.3025-3033
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    • 2016
  • Chronic myeloid leukaemia (CML) is a clonal myeloproliferative hematopoietic stem cell disorder. Deregulated BCR-ABL fusion tyrosine kinase activity is the main cause of CML disease pathogenesis, making BCR-ABL an ideal target for inhibition. Current tyrosine kinase inhibitors (TKIs) designed to inhibit BCR-ABL oncoprotein activity, have completely transformed the prognosis of CML. Interruption of TKI treatment leads to minimal residual disease reside (MRD), thought to reside in TKI-insensitive leukaemia stem cells which remain a potential reservoir for disease relapse. This highlights the need to develop new therapeutic strategies for CML either as small molecule master TKIs or phytopharmaceuticals derived from nature to achieve chronic molecular remission. This review outlines the past, present and future therapeutic approaches for CML including coverage of relevant mechanisms, whether ABL dependent or independent, and epigenetic factors responsible for developing resistance against TKIs. Appearance of mutant clones along the course of therapy either pre-existing or induced due to therapy is still a challenge for the clinician. A proposed in-vitro model of generating colony forming units from CML stem cells derived from diagnostic samples seems to be achievable in the era of high throughput technology which can take care of single cell genomic profiling.

Asynchronous Guidance Filter Design Based on Strapdown Seeker and INS Information (스트랩다운 탐색기 및 INS 정보를 이용한 비동기 유도필터 설계)

  • Park, Jang-Seong;Kim, Yun-young;Park, Sanghyuk;Kim, Yoon-Hwan
    • Journal of the Korean Society for Aeronautical & Space Sciences
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    • v.48 no.11
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    • pp.873-880
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    • 2020
  • In this paper, we propose a guidance filter to estimate line of sight rate with strapdown seeker measurements and INS(Inertial Navigation System) information. The measurements of proposed guidance filter consisted of the LOS(Line of Sight) and relative position that can be calculated with the seeker's measurements, INS information and known target position, also the filter is based on an asynchronous filter to use outputs of the two sensors that are out of synchronous and period. Through the proposed filter, we can reduce the effect on parasitic loop that can be caused by using large time delay seeker and improve the estimation performance.

Conformational Switch and Functional Regulation of Proteins (단백질의 구조 전환과 기능 조절)

  • Yu, Myeong-Hee
    • Proceedings of the Korean Institute of Electrical and Electronic Material Engineers Conference
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    • 2001.11b
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    • pp.3-6
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    • 2001
  • In common globular proteins, the native form is in its most stable state. However, the native form of inhibitory serpins (serine protease inhibitors) and some viral membrane fusion proteins is in a metastable state. Metastability in these proteins is critical to their biological functions. Our previous studies revealed that unusual interactions, such as side-chain overpacking, buried polar groups, surface hydrophobic pockets, and internal cavities are the structural basis of the native metastability. To understand the mechanism by which these structural defects regulate protein functions, cavity-filling mutations of a 1-antitrypsin, a prototype serpin, were characterized. Increasing conformational stability is correlated with decreasing inhibitory activity. Moreover, the activity loss appears to correlate with the decrease in the rate of the conformational switch during complex formation with a target protease. We also increased the stability of a 1-antitrypsin greatly via combining various stabilizing single amino acid substitutions that were distributed throughout the molecule. The results showed that a substantial increase of stability, over 13 kcal/mol, affected the inhibitory activity with a correlation of 11% activity loss per kcal/mol. The results strongly suggest that the native metastability of proteins is indeed a structural design that regulates protein functions and that the native strain of a 1-antitrypsin distributed throughout the molecule regulates the inhibitory function in a concerted manner.

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Conformational Switch and Functional Regulation of Proteins (단백질의 구조 전환과 기능 조절)

  • 유명희
    • Electrical & Electronic Materials
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    • v.14 no.12
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    • pp.3-6
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    • 2001
  • In common globular proteins, the native form is n its most stable state. However, the native form of inhibitory serpins (serine protease inhibitors) and some viral membrane fusion proteins is in a metastable state. Metastability in these proteins is critical to their biological functions. Our previous studies revealed that unusual interactions, such as side-chain overpacking, buried polar groups, surface hydrophobic pockets, ad internal cavities are the structural basis of the native metastability. To understand the mechanism by which these structural defects regulate protein functions, cavity-filling mutations of $\alpha$1-antitrypsin, a prototype serpin, were characterized. Increasing conformational stability is correlated with decreasing inhibitory activity. Moreover, the activity loss appears to correlate with the decrease in the rate of the conformational switch during complex formation with a target protease. We also increased the stability of $\alpha$1-antitrypsin greatly via combining various stabilizing single amino acid substitutions that were distributed throughout the molecule. The results showed that a substantial increase of stability, over 13 kcal/mol, affected the inhibitory activity with a correlation of 11% activity loss per kcal/mol. The results strongly suggest that the native metastability of proteins is indeed a structural design that regulates protein functions and that the native strain of $\alpha$1-antitrypsin distributed throughout the molecule regulates the inhibitory function in a concerted manner.

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3D Orientation and Position Tracking System of Surgical Instrument with Optical Tracker and Internal Vision Sensor (광추적기와 내부 비전센서를 이용한 수술도구의 3차원 자세 및 위치 추적 시스템)

  • Joe, Young Jin;Oh, Hyun Min;Kim, Min Young
    • Journal of Institute of Control, Robotics and Systems
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    • v.22 no.8
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    • pp.579-584
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    • 2016
  • When surgical instruments are tracked in an image-guided surgical navigation system, a stereo vision system with high accuracy is generally used, which is called optical tracker. However, this optical tracker has the disadvantage that a line-of-sight between the tracker and surgical instrument must be maintained. Therefore, to complement the disadvantage of optical tracking systems, an internal vision sensor is attached to a surgical instrument in this paper. Monitoring the target marker pattern attached on patient with this vision sensor, this surgical instrument is possible to be tracked even when the line-of-sight of the optical tracker is occluded. To verify the system's effectiveness, a series of basic experiments is carried out. Lastly, an integration experiment is conducted. The experimental results show that rotational error is bounded to max $1.32^{\circ}$ and mean $0.35^{\circ}$, and translation error is in max 1.72mm and mean 0.58mm. Finally, it is confirmed that the proposed tool tracking method using an internal vision sensor is useful and effective to overcome the occlusion problem of the optical tracker.