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NDP Kinases Suppressed Bax-Dependent Apoptosis in Yeast System

  • K. C. Hwang;D. W. Ok;D. N. Kwon;H. K. Shin;Kim, J. H.
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.52-52
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    • 2001
  • Many nucleoside diphosphate (NDP) kinases are ubiquitous enzymes responsible for the exchange of ${\gamma}$-phosphates between tri- and diphosphonucleosides. The catalytic Many nucleoside diphosphate (NDP) kinases are ubiquitous enzymes responsible for the exchange of ${\gamma}$-phosphates between tri- and diphosphonucleosides. The catalytic reaction follows a ping-pong mechanism in which the enzyme is transiently phosphorylated on a histidine residue conserved in all nucleoside diphosphate kinases. Beside their role in nucleotide synthesis, these enzymes present additional functions, possibly independent of catalysis, in processes such as differentiation, cell growth, tumor progression, metastasis and development. To clone murine nm23-M5, several expressed sequence tags (ESTs) of the GenBank data base, selected according to their homology to nm23-H5 cDNA, reconstituted a complete open reading frame (GenBank AF222750). To test whether murine NDPKs (1, 2, 3, 4, 5, and 6) can inhibit Bax-mediated toxicity in yeast, co-transformation was performed respectively. The yeast S.cerevisiae was transformed with a copy expression plasmid containing the histidine selection marker and expressing murine Bax under the control of a galactose-inducible promoter. Several clones were selected and found to be growth inhibited when Bax expression was induced with galactose. A representative clone was transformed again with a copy expression plasmid containing the tryptophane selection marker and expressing either murine Bcl-xL or NDPK under the control of a galactose-inducible promoter. Several subclones of the double-transformants were selected and characterized. The ability of Bcl-xL and NDPKs to suppress Bax-mediated toxicity was determined by growing yeast cells overnight in galactose media and spot-testing on galactose plates starting with an equal number of yeast cells as determined by taking the OD$_{600}$. Ten-fold serial dilutions were used in the spot-test. Plates were grown at 3$0^{\circ}C$ for 2-3 days. All murine NDPKs suppressed Bax dependent apoptosis. Futher study will be peformed whether Bax-toxicity inhibition was caused by NDP kinase activity or additional function.n.

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Genenation of structural diversity in polyketides by combinatorial biosynthesis of polyketides: Part I. Generation of multiple bioactive macrolides by hybrid modular polyketide synthases in Streptomyces venezuelae, Part II. Production of novel rifamycins by combinatorial biosynthesis

  • Yoon, Yeo-Joon
    • Proceedings of the Korean Society for Applied Microbiology Conference
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    • 2002.10a
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    • pp.18-25
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    • 2002
  • The pikromycin biosynthetic system in Streptomyces venezuleae is unique for its ability to produce two groups of antibiotics that include the 12-membered ring macrolides methymycin and neomethymycin, and the 14-membered ring macrolides narbomycin and pikromycin. The metabolic pathway also contains two post polyketide-modification enzymes, a glycosyltransferase and P450 hydroxylase that have unusually broad substrate specificities. In order to explore further the substrate flexibility of these enzymes a series of hybrid polyketide synthases were constructed and their metabolic products characterized. The plasmid-based replacement of the multifunctional protein subunits of the pikromycin PKS in S. venezuelae by the corresponding subunits from heterologous modular PKSs resulted in recombinant strains that produce both 12- and 14-membered ring macrolactones with predicted structural alterations. In all cases, novel macrolactones were produced and further modified by the DesVII glycosyltransferase and PikC hydroxylase leading to biologically active macrolide structures. These results demonstrate that hybrid PKSs in S. venezuelae can produce a multiplicity of new macrolactones that are modified further by the highly flexible DesVII glycosyltransferase and PikC hydroxylase tailoring enzymes. This work demonstrates the unique capacity of the S. venezuelae pikromycin pathway to expand the toolbox of combinatorial biosynthesis and to accelerate the creation of novel biologically active natural products. The polyketide backbone of rifamycin B is assembled through successive condensation and ${\beta}$-carbonyl processing of the extender units by the modular rifamycin PKS. The eighth module, in the RifD protein, contains nonfunctional DH domain and functional KR domain, which specify the reduction of the ${\beta}$-carbonyl group resulting in the C-21 bydroxyl of rifamycin B. A four amino acid substitution and one amino acid deletion were introduced in the putative NADPH binding motif in the proposed KR domain encoded by rifD. This strategy of mutation was based on the amino acid sequences of the corresponding motif of the KR domain of module 3 in the RifA protein, which is believed dysfunctional, so as to introduce a minimum alteration and retain the reading frame intact, yet ensure loss of function. The resulting strain produces linear polyketides, from tetraketide to octaketide, which are also produced by a rifD disrupted mutant as a consequence of premature termination of polyketide assembly. Much of the structural diversity within the polyketide superfamily of natural products is due to the ability of PKSs to vary the reduction level of every other alternate carbon atom in the backbone. Thus, the ability to introduce heterologous reductive segments such as ketoreductase (KR), dehydratase (DH), and enoylreductase (ER) into modules that naturally lack these activities would increase the power of the combinatorial biosynthetic toolbox. The dehydratase domain of module 7 of the rifamycin PKS, which is predicted to be nonfunctional in view of the sequence of the apparent active site, was replaced with its functional homolog from module 7 of rapamycin-producing polyketide synthase. The resulting mutant strain behaved like a rifC disrupted mutant, i.e., it accumulated the heptaketide intermediate and its precursors. This result points out a major difficulty we have encountered with all the Amycolatopsis mediterranei strain containing hybrid polyketide synthases: all the engineered strains prepared so far accumulate a plethora of products derived from the polyketide chain assembly intermediates as major products instead of just analogs of rifamycin B or its ansamycin precursors.

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Qualitative Evaluation of 2D Dosimetry System for Helical Tomotherapy (2차원 토모테라피 선량측정시스템의 정성적 평가)

  • Ma, Sun Young;Jeung, Tae Sig;Shim, Jang Bo;Lim, Sangwook
    • Progress in Medical Physics
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    • v.25 no.4
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    • pp.193-198
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    • 2014
  • The purpose of this study is to see the feasibility of the newly developed 2D dosimetry system using phosphor screen for helical tomotherapy. The cylindrical water phantom was fabricated with phosphor screen to emit the visible light during irradiation. There are three types of virtual target, one is one spot target, another is C-shaped target, and the other is multiple targets. Each target was planned to be treated at 10 Gy by treatment planning system (TPS) of tomotherapy. The cylindrical phantom was placed on the tomotherapy table and irradiated as calculations of the TPS. Every frame which acquired by CCD camera was integrated and the doses were calculated in pixel by pixel. The dose distributions from the fluorescent images were compared with the calculated dose distribution from the TPS. The discrepancies were evaluated as gamma index for each treatment. The curve for dose rate versus pixel value was not saturated until 900 MU/min. The 2D dosimetry using the phosphor screen and the CCD camera is respected to be useful to verify the dose distribution of the tomotherapy if the linearity correction of the phosphor screen improved.

Setup Verification in Stereotactic Radiotherapy Using Digitally Reconstructed Radiograph (DRR) (디지털화재구성사진(Digitally Reconstructed Radiograph)을 이용한 정위방사선수술 및 치료의 치료위치 확인)

  • Cho, Byung-Chul;Oh, Do-Hoon;Bae, Hoon-Sik
    • Radiation Oncology Journal
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    • v.17 no.1
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    • pp.84-88
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    • 1999
  • Purpose :To develop a method for verifying a treatment setup in stereotactic radiotherapy by ma- tching portal images to DRRs. Materials and Methods : Four pairs of orthogonal portal images of one patient immobilized by a thermoplastic mask frame for fractionated stereotactic radiotherapy were compared with DRRs. Portal images are obtained in AP (anteriorfposterior) and lateral directions with a target localizer box containing fiducial markers attached to a stereotactic frame. DRRs superimposed over a planned iso-center and fiducial markers are printed out on transparent films. And then, they were overlaid over onhogonal penal images by matching anatomical structures. From three different kind of objects (isgcenter, fiducial markers, anatomical structure) on DRRs and portal images, the displacement error between anatomical structure and isocenters (overall setup error), the displacement error between anatomical structure and fiducial markers (irnrnobiliBation error), and the displacement error between fiducial markers and isocenters (localization error) were measured. Results : Localization error were 1.5$\pm$0.3 mm (AP), 0.9$\pm$0.3 mm (lateral), and immobilization errors were 1.9$\pm$0.5 mm (AP), 1.9$\pm$0.4 mm (lateral). In addition, overall setup errors were 1.0$\pm$0.9 mm (AP), 1.3$\pm$0.4 mm (lateral). From these orthogonal displacement errors, maximum 3D displacement errors($\sqrt{(\DeltaAP)^{2}+(\DeltaLat)^{2}$)) were found to be 1.7$\pm$0.4 mm for localization, 2.0$\pm$0.6 mm for immobilization, and 2.3$\pm$0.7 mm for overall treatment setup. Conclusion : By comparing orthogonal portal images with DRRs, we find out that it is possible to verify treatment setup directly in stereotactic radiotherapy.

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Cloning, Sequencing and Expression of the Gene Encoding a Thermostable β-Xylosidase from Paenibacillus sp. DG-22 (Paenibacillus sp. DG-22로부터 열에 안정한 β-xylosidase를 암호화하는 유전자의 클로닝, 염기서열결정 및 발현)

  • Lee, Tae-Hyeong;Lee, Yong-Eok
    • Journal of Life Science
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    • v.17 no.9 s.89
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    • pp.1197-1203
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    • 2007
  • A genomic DNA library of the bacterium Paenibacillus sp. DG-22 was constructed and the ${\beta}-xylosi-dase-positive$ clones were identified using the fluorogenic substrate $4-methylumbelliferyl-{\beta}-D-xylopyr-anoside$ $({\beta}MUX)$. A recombinant plasmid was isolated from the clone and 4.3-kb inserted DNA was sequenced. The ${\beta}-xylosidase$ gene (xylA) was comprised of a 2,106 bp open reading frame (ORF) en-coding 701 amino acids with a molecular weight of 78,710 dalton and a pI of 5.0. The deduced amino acid sequence of the xylA gene product had significant similarity with ${\beta}-xylosidases$ classified into family 52 of glycosyl hydrolases. The xylA gene was subcloned into the pQE60 expression vector to fuse with six histidine-tag. The recombinant ${\beta}-xylosidase$ $(XylA-H_6)$ was purified to homogeneity by heat-treatment and immobilized metal affinity chromatography. The pH and temperature optima of the $XylA-H_6$ enzyme were pH 5.5-6.0 and $60^{\circ}C$, respectively.

A Study for Relationship Between the Femininity and the Nursing Image of Nursing Students (일부 간호학생의 여성성과 간호상과의 상관관계)

  • 성현란
    • Journal of Korean Academy of Nursing
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    • v.8 no.2
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    • pp.39-49
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    • 1978
  • The present research set out to further validate the results that Stromborg (1976) obtained in which a correlation (r= .22, p < .001) was found between the femininity of woman nursing students and their image of nursing. In addition the experimenter set out to elucidate any possible differences in the image of nursing according to (a) religion, (b) the motivation for choosing nursing as well as, (c) the particular area in which the students hoped to work after graduating. The research was conducted by presenting questionnaire type scales including a femininity scale and a nursing image scale, to 301 graduating students attending 4 Three-year nursing schools located within Seoul from November 21st, 1978 to November 30th. The results: 1. There was clearly no relation between femininity and the image of nursing as such (r= .01, p > .43). But upon analyzing the results along five subdimensions of the nursing image variable it was discovered that : (a) if femininity is low, the nursing student tend to view the educational preparation for becoming a nurse in a more positive light (r= -.10, p < .05), and (b) if femininity is high, the students tend to perceive their social role with a positive frame of mind (r= .15, p < .005). 2. There was a significant difference found in the image of nursing as related to the religion of the nursing students, The means ranked as follows (high scores indicate a positive image of the nursing profession). Catholic > Protestant > no religion > Buddhist 3, The difference in the nursing image according to motivation for sellecting the profession emerged as quite significant (F=6.92, p < .001). The rank of the means when the subject group is divided along the dimension of motive is as follows (ranking begins with the most positive nursing image): (a) The profession itself seemed good; (b) after marriage one can do a social type of activity; (c) significant others’advice: (d) the profession matches their high school grade record; (e) it matches the economic situation of the student; (f)no particular reason for their choice: (g) it's easy to get a nursing job after graduation. 4. The particular area of nursing one intended to go into after graduation varied significantly in terms of the image of nursing (F=3.03, p < .005). The means when ranked starting with those having the most positive image of the profession: (a) nurses helping deliveries; (b) hospital nurses: (c) nursing education; (d) school nurses: (e) government nurse supervisors: (f)public health nurses: (g) (students not going into any profession); (h) (students not going into the nursing profession but into some other area).

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Optical Microphone Incorporating a Reflective Micromirror and a Dual-core Collimator (반사형 마이크로미러와 듀얼 코어 클리메이터를 이용한 광 마이크로폰)

  • Song, Ju-Han;Kim, Do-Hwan;Gu, Hyun-Mo;Park, Hyun-Jung;Lee, Sang-Shin;Cho, Il-Joo
    • Korean Journal of Optics and Photonics
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    • v.17 no.1
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    • pp.94-98
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    • 2006
  • An optical microphone based on a dual-core fiber collimator and a membrane type micromirror serving as an optical head and a reflective diaphragm respectively was implemented. The micromirror diaphragm is suspended by a thin silicon bar linked with a frame, thus it is subject to a displacement induced by acoustic waves. The optical head incorporating two collimators integrated in a single housing gives light to and receives it from the diaphragm, rendering the optical microphone structure simple and compact. This dual-core collimator having a slowing varying beam profile facilitates the initial alignment of the optical head with the diaphragm, especially the distance between them. For the assembled microphone, the static characteristics were investigated tofind the operation point defined as the optimum distance between the head and the diaphragm, and a frequency response with a variation of about $\pm$5 dB for the range of up to 3kHz was achieved.

A Frame-based Coding Mode Decision for Temporally Active Video Sequence in Distributed Video Coding (분산비디오부호화에서 동적비디오에 적합한 프레임별 모드 결정)

  • Hoangvan, Xiem;Park, Jong-Bin;Shim, Hiuk-Jae;Jeon, Byeung-Woo
    • Journal of Broadcast Engineering
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    • v.16 no.3
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    • pp.510-519
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    • 2011
  • Intra mode decision is a useful coding tool in Distributed Video Coding (DVC) for improving DVC coding efficiency for video sequences having fast motion. A major limitation associated with the existing intra mode decision methods, however, is that its efficiency highly depends on user-specified thresholds or modeling parameters. This paper proposes an entropy-based method to address this problem. The probabilities of intra and Wyner?Ziv (WZ) modes are determined firstly by examining correlation of pixels in spatial and temporal directions. Based on these probabilities, entropy of the intra and the WZ modes are computed. A comparison based on the entropy values decides a coding mode between intra coding and WZ coding without relying on any user-specified thresholds or modeling parameters. Experimental results show its superior rate-distortion performance of improvements of PSNR up to 2 dB against a conventional Wyner?Ziv coding without intra mode decision. Furthermore, since the proposed method does not require any thresholds or modeling parameters from users, it is very attractive for real life applications.

Uniform Posture Map Algorithm to Generate Natural Motion Transitions in Real-time (자연스러운 실시간 동작 전이 생성을 위한 균등 자세 지도 알고리즘)

  • Lee, Bum-Ro;Chung, Chin-Hyun
    • Journal of KIISE:Computing Practices and Letters
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    • v.7 no.6
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    • pp.549-558
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    • 2001
  • It is important to reuse existing motion capture data for reduction of the animation producing cost as well as efficiency of producing process. Because its motion curve has no control point, however, it is difficult to modify the captured data interactively. The motion transition is a useful method to reuse the existing motion data. It generates a seamless intermediate motion with two short motion sequences. In this paper, Uniform Posture Map (UPM) algorithm is proposed to perform the motion transition. Since the UPM is organized through quantization of various postures with an unsupervised learning algorithm, it places the output neurons with similar posture in adjacent position. Using this property, an intermediate posture of two active postures is generated; the generating posture is used as a key-frame to make an interpolating motion. The UPM algorithm needs much less computational cost, in comparison with other motion transition algorithms. It provides a control parameter; an animator could control the motion simply by adjusting the parameter. These merits of the UPM make an animator to produce the animation interactively. The UPM algorithm prevents from generating an unreal posture in learning phase. It not only makes more realistic motion curves, but also contributes to making more natural motions. The motion transition algorithm proposed in this paper could be applied to the various fields such as real time 3D games, virtual reality applications, web 3D applications, and etc.

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A Study on the automatic vehicle monitoring system based on computer vision technology (컴퓨터 비전 기술을 기반으로 한 자동 차량 감시 시스템 연구)

  • Cheong, Ha-Young;Choi, Chong-Hwan;Choi, Young-Gyu;Kim, Hyon-Yul;Kim, Tae-Woo
    • The Journal of Korea Institute of Information, Electronics, and Communication Technology
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    • v.10 no.2
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    • pp.133-140
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    • 2017
  • In this paper, we has proposed an automatic vehicle monitoring system based on computer vision technology. The real-time display system has displayed a system that can be performed in automatic monitoring and control while meeting the essential requirements of ITS. Another advantage has that for a powerful vehicle tracking, the main obstacle handing system, which has the shadow tracking of moving objects. In order to obtain all kinds of information from the tracked vehicle image, the vehicle must be clearly displayed on the surveillance screen. Over time, it's necessary to precisely control the vehicle, and a three-dimensional model-based approach has been also necessary. In general, each type of vehicle has represented by the skeleton of the object or wire frame model, and the trajectory of the vehicle can be measured with high precision in a 3D-based manner even if the system has not running in real time. In this paper, we has applied on segmentation method to vehicle, background, and shadow. The validity of the low level vehicle control tracker was also detected through speed tracking of the speeding car. In conclusion, we intended to improve the improved tracking method in the tracking control system and to develop the highway monitoring and control system.