• Title/Summary/Keyword: Time-of-flight(TOF)

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Cloning, Over-expression, and Characterization of YjgA, a Novel ppGpp-binding Protein

  • Gnanasekaran, Gopalsamy;Pan, SangO;Jung, Wontae;Jeong, Kwangjoon;Jeong, Jae-Ho;Rhee, Joon Haeng;Choy, Hyon E.;Jung, Che-Hun
    • Bulletin of the Korean Chemical Society
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    • v.34 no.8
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    • pp.2419-2424
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    • 2013
  • Guanosine-5'-diphosphate 3'-diphosphate (ppGpp) serves as alarmone in bacterial stringent responses. In this study, an affinity column was constructed by immobilizing ppGpp to NHS-Sepharose for isolating ppGpp-binding proteins. A novel ppGpp-binding protein, YjgA, was isolated and characterized by MALDI-TOF MS (matrix-assisted laser desorption ionization-time-of-flight mass spectrometry) coupled with two-dimensional gel electrophoresis. YjgA and truncated forms of YjgA were cloned and over-expressed in BL21 (DE3). The binding affinity of YjgA to ppGpp was determined by equilibrium dialysis. The interaction of YjgA with ppGpp was very specific, considering that the dissociation constant of YjgA with ppGpp was measured as $5.2{\pm}2.0{\mu}M$, while the affinities to GTP and GDP were about 60 and 30 times weaker than ppGpp. Expression of yjgA gene in Escherichia coli K-12 MG1655 was examined by reverse transcription polymerase chain reaction (RT-PCR). RT-PCR results revealed that yjgA was expressed from early to late stationary phase. The yjgA deletion mutant exhibited decreased cell number at stationary phase compared to parent strain and the over-expression of YjgA increased the cell number. These results suggested that YjgA might stimulate cell division under stationary phase. In most prokaryotic genome, about half of the protein candidates are hypothetical, that are expected to be expressed but there is no experimental report on their functions. The approach utilized in this study may serve as an effective mean to probe the functions of hypothetical proteins.

Monoclonal antibody production for CP4 EPSPS detection assays (CP4 EPSPS 검출을 위한 단클론 항체 생산)

  • A-Mi Yoon;Il Ryong Kim;Wonkyun Choi
    • Korean Journal of Environmental Biology
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    • v.39 no.4
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    • pp.445-451
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    • 2021
  • In this study, we described the production of an antibody to living modified organisms (LMOs) containing the gene encoding for 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) from Agrobacterium tumefaciens strain CP4 EPSPS provides resistance to the herbicide glyphosate (N- (phosphonomethyl) glycine). These LMOs were approved and have recently been used in the feed, food production, and processing industries in South Korea. Highly efficient monoclonal antibody (mAb) production is crucial for developing assays that enable the proper detection and quantification of the CP4 EPSPS protein in LMOs. This study describes the purification and characterization of recombinant CP4 EPSPS protein in E. coli BL21 (DE3) based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrixassisted laser desorption/ionization time-of-flight mass spectrometry. The production of mAbs was undertaken based on the standard operating procedure of Abclon, Inc.(South Korea), and the purity of the mAbs was assessed using SDS-PAGE. The following five mAb clones were produced: 2F2, 4B9, 6C11, 10A9, and 10G9. To verify the efficiency and specificity of the five developed mAbs, we performed Western blotting analysis using the LM (living modified) cotton crude extracts. All mAbs could detect the CP4 EPSPS protein in the LM cotton traits MON1445 and MON88913 with high specificity, but not in any other LM cottons or non-LM cottons. These data indicate that these five mAbs to CP4 EPSPS could be successfully used for the further development of antibody-based detection methods to target CP4 EPSPS protein in LMOs.

Proteomics of Protein Expression Profiling in Tissues with Different Radiosensitivity (Proteomics를 이용한 마우스 조직에서의 방사선 감수성 조절 단백질의 탐색)

  • An, Jeung-Hee;Kim, Ji-Young;Seong, Jin-Sil
    • Radiation Oncology Journal
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    • v.22 no.4
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    • pp.298-306
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    • 2004
  • Purpose: The purpose of this study was to identify Radiosensitivity of proteins in tissues with different radiosensitivity. Materials and Methods: C3H/HeJ mice were exposed to 10 Gy. The mice were sacrifiud 8 hrs after radiation. Their spleen and liver tissues were collected and analyzed histologicaly for apoptosis. The expressions of radiosusceptibillty protein were analyzed by 2-dimensional electrophoresis and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Resilts: The Peak of apoptosis levels were $35.3{\pm}1.7{\%}$ in spleen and $0.6{\pm}0.2{\%}$ in liver at 8 hrs after radiation. Liver, radioresistant tissues, showed that the levels of ROS metabolism related to proteins such as cytochromm c, glutathione S transferase, NADH dehydrogenase, riken cDNA and peroxiredoxin Vl increased after radiation. The expression of cytochrome c increased significantly in spleen and liver tissues after radiation. In spleen, radiosensitivity tissue, the identified proteins showed a significantly quantitative alteration following radiation. It was categorized as signal transduction, apoptosis, cytokine, Ca signal related protein, stress-related protein, cytoskeletal regulation, ROS metabolism, and others. Conclusion: Differences of radiation-induced apoptosis by tissues specifted were coupled with the induction of related radiosensitivity and radioresistant proteins. The result suggests that apoptosis relate protein and redox proteins play important roles in this radiosusceptibility.

Characteristics of Coagulase-negative Staphylococci Isolates from Dental Clinic Environments in Busan, Korea (부산지역 치과환경에서 분리된 coagulase-negative staphylococci의 특성)

  • Jung, Hye-In;Jung, So Young;Park, Indal;Bae, Il Kwon
    • Journal of Life Science
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    • v.26 no.2
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    • pp.220-225
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    • 2016
  • Coagulase-negative staphylococci (CNS) have recently become the bacteria most frequently found in clinical infections. The aim of this study was to investigate the prevalence, antimicrobial susceptibilities, and molecular characteristics of CNS isolates from dental clinic environments in Busan, Korea. One hundred and fifty-four samples were collected from 10 dental clinics and dental hospitals in Busan from December 2014 to January 2015. Species were identified by matrix-assisted laser desorption/ionization–time-of-flight. Antimicrobial susceptibility was determined by disk diffusion methods. A polymerase chain reaction was performed to detect mecA, mupA gene, and SCCmec types. Of the 154 samples, 10(6.5%) isolates were identified as CNS (5 Staphylococcus epidermidis, 2 Staphylococcus capitis, 2 Staphylococcus, and 1 Staphylococcus haemolyticus). Among the 10 isolates, 6 were resistant to penicillin, 5 were resistant to gentamicin, 3 were resistant to tetracycline, and 2 were resistant to cefoxitin and erythromycin. However, clindamycin, ciprofloxacin, teicoplanin, and trimethoprim-sulfamethoxazole resistant isolates were not present. Genes encoding mecA were detected in 4 (2 S. warneri and 2 S. haemolyticus) isolates, and mupA in 1 (S. epidermidis) isolate. One methicillin-resistant CNS (S. warneri) isolate was determined as being of the SCCmec type I. It is concluded that CNS resistant to various antimicrobial agents was widely distributed in dental clinic environments in Korea.

Protein Profile in Corpus Luteum during Pregnancy in Korean Native Cows

  • Chung, H.J.;Kim, K.W.;Han, D.W.;Lee, H.C.;Yang, B.C.;Chung, H.K.;Shim, M.R.;Choi, M.S.;Jo, E.B.;Jo, Y.M.;Oh, M.Y.;Jo, S.J.;Hong, S.K.;Park, J.K.;Chang, W.K.
    • Asian-Australasian Journal of Animal Sciences
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    • v.25 no.11
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    • pp.1540-1545
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    • 2012
  • Steroidogenesis requires coordination of the anabolic and catabolic pathways of lipid metabolism, but the profile of proteins associated with progesterone synthesis in cyclic and pregnant corpus luteum (CL) is not well-known in cattle. In Experiment 1, plasma progesterone level was monitored in cyclic cows (n = 5) and pregnant cows (n = 6; until d-90). A significant decline in the plasma progesterone level occurred at d-19 of cyclic cows. Progesterone level in abbatoir-derived luteal tissues was also determined at d 1 to 5, 6 to 13 and 14 to 20 of cyclic cows, and d-60 and -90 of pregnant cows (n = 5 each). Progesterone level in d-60 CL was not different from those in d 6 to 13 CL and d-90 CL, although the difference between d 6 to 13 and d-90 was significant. In Experiment 2, protein expression pattern in CL at d-90 (n = 4) was compared with that in CL of cyclic cows at d 6 to 13 (n = 5). Significant changes in the level of protein expression were detected in 32 protein spots by two-dimensional polyacrylamide gel electrophoresis (2-DE), and 23 of them were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Six proteins were found only in pregnant CL, while the other 17 proteins were found only in cyclic CL. Among the above 6 proteins, vimentin which is involved in the regulation of post-implantation development was included. Thus, the protein expression pattern in CL was disorientated from cyclic luteal phase to mid pregnancy, and alterations in specific CL protein expression may contribute to the maintenance of pregnancy in Korean native cows.

3-Dimensional LADAR Optical Detector Development in Geiger Mode Operation (Geiger Mode로 동작하는 3차원 LADAR 광수신기 개발)

  • Choi, Soon-Gyu;Shin, Jung-Hwan;Kang, Sang-Gu;Hong, Jung-Ho;Kwon, Yong-Joon;Kang, Eung-Cheol;Lee, Chang-Jae
    • Korean Journal of Optics and Photonics
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    • v.24 no.4
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    • pp.176-183
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    • 2013
  • In this paper, we report the design, fabrication and characterization of the 3-Dimensional optical receiver for a Laser Detection And Ranging (LADAR) system. The optical receiver is composed of three parts; $16{\pm}16$ Geiger Mode InGaAs Avalanche Photodiode (APD) array device operated at 1560 nm wavelength, Read Out Integrated Circuit (ROIC) measuring the Time-Of-Flight (TOF) of the return signal reflected from target objects, a package and cooler maintaining the proper operational condition of the detector and control electronics. We can confirm that the LADAR system can detect the signal from a target up to 1.2 km away, and it showed low Dark Count Rate (DCR) of less than 140 kHz, and higher than 28%-Photon Detection Efficiency (PDE). This is considered to be the best performance of the $16{\pm}16$ FPA APD optical receiver for a LADAR system.

Changes in Total Cerebral Blood Flow with Aging, Parenchymal Volume Changes, and Vascular Abnormalities: a Two-dimensional Phase-Contrast MRI Study (나이와 뇌실질부피 변화 및 혈관이상에 따른 총뇌혈류량 변화: 이차원 위상대조 자기공명영상을 이용한 연구)

  • Liu Haiying;Shin Tae-Beom;Youn Seong-Kuk;Oh Jong-Yong;Lee Young-Il;Choi Sun-Seob
    • Investigative Magnetic Resonance Imaging
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    • v.8 no.1
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    • pp.17-23
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    • 2004
  • Purpose : To evaluate changes in total cerebral blood flow (tCBF) with aging, parenchymal volume changes and vascular abnormalities, using 2 dimensional (D) phase-contrast magnetic resonance imaging (PC MRI). Materials and Methods : Routine brain MRI including T2 weighted image, time-of-flight (TOF) MR Angiography (MRA) and 2D PC MRI were performed in 73 individuals, including 12 volunteers. Normal subjects (12 volunteers, and 21 individuals with normal MRI and normal MRA) were classified into groups according to age (18-29, 30-49 and 50-66 years). For the group with abnormalities in brain MRIs, cerebral parenchymal volume changes were scored according to the T2 weighted images, and atherosclerotic changes were scored according to the MRA findings. Abnormal groups were classified into 4 groups: (i) mild reduction in volume, (ii) marked reduction in volume by parenchymal volume and atherosclerotic changes, and (iii) increased volume and (iv) Moya-moya disease. Volumetric flow was measured at the internal carotid artery (ICA) and vertebral artery bilaterally using the velocity-flow diagrams from PC MRI, and combined 4 vessel flows and tCBF were compared among all the groups. Results : The age-specific distribution of tCBFs in normal subjects were as follows: $12.0{\pm}2.1ml/sec$ in 18-29 years group, $11.8{\pm}1.9ml/sec$ in 30-49 years group, $10.9{\pm}2.2ml/sec$ in 50-66 years group. The distribution of tCBFs in the different subsets of the abnormal population were as follows: $9.5{\pm}2.5ml/sec$ in the group with mild reduction in volume, $7.6{\pm}2.0ml/sec$ in the group with marked reduction in volume, and $7.3{\pm}1.2ml/sec$ and $7.0{\pm}1.1ml/sec$ in the increased parenchymal volume and Moya-moya disease groups respectively. Conclusion : Total cerebral blood flow decreases with increasing age with a concomitant reduction in parenchymal volumes and increasing atherosclerotic changes. It is also reduced in the presence of increased parenchymal volume and Moya-moya disease.2D PC MRI can be used as a tool to evaluate tCBF with aging and in the presence of various conditions that can affect parenchymal volume and cerebral vasculature.

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Chemical compositions of different cultivars of astringent persimmon (Diospyros kaki Thunb.) and the effects of maturity (떫은감(Diospyros kaki Thunb.) 품종 및 수확시기에 따른 영양 성분 변화)

  • Yoo, Seul Ki;Kim, Jong Min;Park, Seon Kyeong;Kang, Jin Yong;Han, Hye Ju;Park, Hyo Won;Kim, Chul-Woo;Lee, Uk;Heo, Ho Jin
    • Korean Journal of Food Science and Technology
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    • v.51 no.3
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    • pp.248-257
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    • 2019
  • This study aimed to determine the chemical components of different cultivars (Gabjubaekmok, Sangjudungsi, and Godongsi) of persimmon (Diospyros kaki Thunb.) and the effects of maturity. The detected free sugars were fructose and glucose, and glucose levels tended to increase with maturity. Palmitic acid, oleic acid, linoleic acid and stearic acid, and potassium were major components. In the amino acid component analysis, the following were usually present in the following month: glutamic acid in Gabjubaekmok in July, lysine in Sangjudungsi in August, and threonine in Godongsi in October. Vitamin C tended to increase with aging, and fruit harvested in July had the highest total phenolic and flavonoid contents. In the metabolite analysis, there were significant differences among cultivars and with maturity. The major physiological compounds were analyzed using ultra performance liquid chromatography quadrupole time-of-flight tandem mass ($UPLC/Q-TOF/MS^2$) and were citric acid and gallic aicd. As maturity progressed, citric acid increased but gallic acid decreased.

Evaluation of TOF MR Angiography and Imaging for the Half Scan Factor of Cerebral Artery (유속신호증강효과의 자기공명혈관조영술을 이용한 뇌혈관검사에서 Half Scan Factor 적용한 영상 평가)

  • Choi, Young Jae;Kweon, Dae Cheol
    • Journal of the Korean Magnetics Society
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    • v.26 no.3
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    • pp.92-98
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    • 2016
  • To aim of this study was to assess the full scan and half scan of imaging with half scan factor. Patients without a cerebral vascular disease (n = 30) and were subject to the full scan half scan, and set a region of interest in the cerebral artery from the three regions (C1, C2, C3) in the range of 7 to 8 mm. MIP (maximum intensity projection) to reconstruct the images in signal strength SNR (signal to noise ration), PSNR (peak signal noise to ratio), RMSE (root mean square error), MAE (mean absolute error) and calculated by paired t-test for use by statistics were analyzed. Scan time was half scan (4 minutes 53 seconds), the full scan (6 minutes 04 seconds). The mean measurement range (7.21 mm) of all the ROI in the brain blood vessel, was the SNR of the first C1 is completely scanned (58.66 dB), half-scan (62.10 dB), a positive correlation ($r^2=0.503$), for the second C2 SNR is completely scanned (70.30 dB), half-scan (74.67 dB) the amount of correlation ($r^2=0.575$), third C3 of a complete scan SNR (70.33 dB), half scan SNR (74.64 dB) in the amount of correlation between the It was analyzed with ($r^2=0.523$). Comparative full scan with half of SNR ($4.75{\pm}0.26dB$), PSNR ($21.87{\pm}0.28dB$), RMSE ($48.88{\pm}1.61$), was calculated as MAE ($25.56{\pm}2.2$). SNR is also applied to examine the half-scans are not many differences in the quality of the two scan methods were not statistically significant in the scan (p-value > .05) image takes less time than a full scan was used.

Phosphate Concentration Dependent Degradation of Biofilm in S. aureus Triggered by Physical Properties (인산염 농도에 따른 물성 변화로 발생하는 황색포도상구균 바이오필름 제거 현상)

  • Song, Sang-Hun;Hwang, Byung Woo;Son, Seong Kil;Kang, Nae-Gyu
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.47 no.4
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    • pp.361-368
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    • 2021
  • The objective of this study was to establish technology for removing bacteria with human- and eco-friendly material. Staphylococcus aureus as an important component for balanced equilibrium among microbiomes, was cultured under various concentrations of phosphate. Experimental observation relating to physical properties was performed in an addition of phosphate buffer. Statistically minimum value of size and hardness using atomic force microscope was observed on the matured biofilm at 5 mM concentration of phosphate. As a result of absorbance for the biofilm tagged with dye, concentration of biofilm was reduced with phophate, too. To identify whether this reduction by phosphate at the 5 mM is caused by counter ion or not, sodium chloride was treated to the biofilm under the same condition. To elucidate components of the biofilm counting analysis of the biofilm using time-of-flight secondary ion mass spectrometry was employed. The secondary ions from the biofilm revealed that alteration of physical properties is consistent to the change of extracellular polymeric substrate (EPS) for the biofilm. Viscoelastic characterization of the biofilm using a controlled shear stress rheometer, where internal change of physical properties could be detected, exhibited a static viscosity and a reduction of elastic modulus at the 5 mM concentration of phosphate. Accordingly, bacteria at the 5 mM concentration of phosphate are attributed to removing the EPS through a reduction of elastic modulus for bacteria. We suggest that the reduction of concentration of biofilm induces dispersion which assists to easily spread its dormitory. In conclusion, it is elucidated that an addition of phosphate causes removal of EPS, and that causes a function of antibiotic.