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Effects of N-/C-Terminal Extra Tags on the Optimal Reaction Conditions, Activity, and Quaternary Structure of Bacillus thuringiensis Glucose 1-Dehydrogenase

  • Hyun, Jeongwoo;Abigail, Maria;Choo, Jin Woo;Ryu, Jin;Kim, Hyung Kwoun
    • Journal of Microbiology and Biotechnology
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    • v.26 no.10
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    • pp.1708-1716
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    • 2016
  • Glucose dehydrogenase (GDH) is an oxidoreductase enzyme and is used as a biocatalyst to regenerate NAD(P)H in reductase-mediated chiral synthesis reactions. In this study, the glucose 1-dehydrogenase B gene (gdhB) was cloned from Bacillus thuringiensis subsp. kurstaki, and wild-type (GDH-BTWT) and His-tagged (GDH-BTN-His, GDH-BTC-His) enzymes were produced in Escherichia coli BL21 (DE3). All enzymes were produced in the soluble forms from E. coli. GDH-BTWT and GDH-BTN-His showed high specific enzymatic activities of 6.6 U/mg and 5.5 U/mg, respectively, whereas GDH-BTC-His showed a very low specific enzymatic activity of 0.020 U/mg. These results suggest that the intact C-terminal carboxyl group is important for GDH-BT activity. GDH-BTWT was stable up to 65℃, whereas GDH-BTN-His and GDH-BTC-His were stable up to 45℃. Gel permeation chromatography showed that GDH-BTWT is a dimer, whereas GDH-BTN-His and GDH-BTC-His are monomeric. These results suggest that the intact N- and C-termini are required for GDH-BT to maintain thermostability and to form its dimer structure. The homology model of the GDH-BTWT single subunit was constructed based on the crystal structure of Bacillus megaterium GDH (PDB ID 3AY6), showing that GDH-BTWT has a Rossmann fold structure with its N- and C-termini located on the subunit surface, which suggests that His-tagging affected the native dimer structure. GDH-BTWT and GDH-BTN-His regenerated NADPH in a yeast reductase-mediated chiral synthesis reaction, suggesting that these enzymes can be used as catalysts in fine-chemical and pharmaceutical industries.

Identification and Characterization of Protein Encoded by orf382 as $\small{L}$-Threonine Dehydrogenase

  • Ma, Fei;Wang, Tianwen;Ma, Xingyuan;Wang, Ping
    • Journal of Microbiology and Biotechnology
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    • v.24 no.6
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    • pp.748-755
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    • 2014
  • In the genome annotation of Escherichia coli MG1655, the orf382 (1,149 bp) is designated as a gene encoding an alcohol dehydrogenase that may be Fe-dependent. In this study, the gene was amplified from the genome by PCR and overexpressed in Escherichia coli BL21(DE3). The recombinant $6{\times}$His-tag protein was then purified and characterized. In an enzymatic assay using different hydroxyl-containing substrates (n-butanol, $\small{L}$-threonine, ethanol, isopropanol, glucose, glycerol, $\small{L}$-serine, lactic acid, citric acid, methanol, or $\small{D}$-threonine), the enzyme showed the highest activity on $\small{L}$-threonine. Characterization of the mutant constructed using gene knockout of the orf382 also implied the function of the enzyme in the metabolism of $\small{L}$-threonine into glycine. Considering the presence of tested substrates in living E. coli cel ls and previous literature, we believed that the suitable nomenclature for the enzyme should be an $\small{L}$-threonine dehydrogenase (LTDH). When using $\small{L}$-threonine as the substrate, the enzyme exhibited the best catalytic performance at $39^{\circ}C$ and pH 9.8 with $NAD^+$ as the cofactor. The determination of the Km values towards $\small{L}$-threonine (Km = $11.29{\mu}M$), ethanol ($222.5{\mu}M$), and n-butanol ($8.02{\mu}M$) also confirmed the enzyme as an LTDH. Furthermore, the LTDH was shown to be an ion-containing protein based on inductively coupled plasma-atomic emission spectrometry with an isoelectronic point of pH 5.4. Moreover, a circular dichroism analysis revealed that the metal ion was structurally and enzymatically essential, as its deprivation remarkably changed the ${\alpha}$-helix percentage (from 12.6% to 6.3%).

Modeling Age-specific Cancer Incidences Using Logistic Growth Equations: Implications for Data Collection

  • Shen, Xing-Rong;Feng, Rui;Chai, Jing;Cheng, Jing;Wang, De-Bin
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.22
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    • pp.9731-9737
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    • 2014
  • Large scale secular registry or surveillance systems have been accumulating vast data that allow mathematical modeling of cancer incidence and mortality rates. Most contemporary models in this regard use time series and APC (age-period-cohort) methods and focus primarily on predicting or analyzing cancer epidemiology with little attention being paid to implications for designing cancer registry, surveillance or evaluation initiatives. This research models age-specific cancer incidence rates using logistic growth equations and explores their performance under different scenarios of data completeness in the hope of deriving clues for reshaping relevant data collection. The study used China Cancer Registry Report 2012 as the data source. It employed 3-parameter logistic growth equations and modeled the age-specific incidence rates of all and the top 10 cancers presented in the registry report. The study performed 3 types of modeling, namely full age-span by fitting, multiple 5-year-segment fitting and single-segment fitting. Measurement of model performance adopted adjusted goodness of fit that combines sum of squred residuals and relative errors. Both model simulation and performance evalation utilized self-developed algorithms programed using C# languade and MS Visual Studio 2008. For models built upon full age-span data, predicted age-specific cancer incidence rates fitted very well with observed values for most (except cervical and breast) cancers with estimated goodness of fit (Rs) being over 0.96. When a given cancer is concerned, the R valuae of the logistic growth model derived using observed data from urban residents was greater than or at least equal to that of the same model built on data from rural people. For models based on multiple-5-year-segment data, the Rs remained fairly high (over 0.89) until 3-fourths of the data segments were excluded. For models using a fixed length single-segment of observed data, the older the age covered by the corresponding data segment, the higher the resulting Rs. Logistic growth models describe age-specific incidence rates perfectly for most cancers and may be used to inform data collection for purposes of monitoring and analyzing cancer epidemic. Helped by appropriate logistic growth equations, the work vomume of contemporary data collection, e.g., cancer registry and surveilance systems, may be reduced substantially.

Comparison of PI and PR Controller Based Current Control Schemes for Single-Phase Grid-Connected PV Inverter (단상 계통 연계형 태양광 인버터에 사용되는 PI 와 PR 전류제어기의 비교 분석)

  • Vu, Trung-Kien;Seong, Se-Jin
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.11 no.8
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    • pp.2968-2974
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    • 2010
  • Nowadays, the PV systems have been focused on the grid connection between the power source and the grid. The PV inverter can be considered as the core of the whole system because of an important role in the grid-interfacing operation. An important issue in the inverter control is the load current regulation. In the literature, Proportional Integral (PI) controller, which is normally used in the current-controlled Voltage Source Inverter (VSI), cannot be a satisfactory controller for an AC system because of the steady-sate error and the poor disturbance rejection, especially in high-frequency range. Compared with conventional PI controller, Proportional Resonant (PR) controller can introduce an infinite gain at the fundamental frequency of the AC source; hence it can achieve the zero steady-state error without requiring the complex transformation and the de-coupling technique. Theoretical analyses of both PI and PR controller are presented and verified by simulation and experiment. Both controller are implemented in a 32-bit fixed-point TMS320F2812 DSP processor and evaluated on a 3kW experimental prototype PV Power Conditioning System (PCS). Simulation and experimental results are shown to verify the controller performances.

Abstracts in Medical Science Journals: An Analysis of Subheadings in Structured Abstracts (의학 저널에서 사용되는 구조적 초록의 소표제들에 관한 분석)

  • Kim, Eungi
    • Journal of Korean Library and Information Science Society
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    • v.47 no.1
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    • pp.199-216
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    • 2016
  • This study aimed to investigate the current uses of subheadings that appear in medical science journal abstracts and to discuss its potential implications for medical science from the perspectives of library and information science. To conduct this study, the following nine sub-fields in medical science were selected: cancer, ethics, genetics, infectious disease, neurology, pediatrics, immunology, psychiatry, and cardiology. Random sample data were drawn based on the years 2010 to 2015 from the PubMed database. This study investigated the extent of the uses of subheadings, variants of subheadings, and common formation of subheadings with the help of a frequency analysis. The specific findings of this study are summarized as the following: 1) more traditional abstracts are used across almost all sub-fields of medical science; 2) on average, 4.1 subheadings were used in the sample dataset; and 3) the most frequently used set of subheadings is OBJECTIVES, METHODS, RESULTS, and CONCLUSIONS. This subheading set appears to be the de facto standard across all medical science journals. The analysis of subheadings in structured abstracts and the issues raised in this study can be beneficial for journal editors and other academics in medical science as well as library and information science.

A Study on the Changes in Regulatory Policy against Large-scale Retail Stores in Japan (일본의 대규모 소매점포 규제 정책 변화에 관한 연구)

  • Kim, Seung-Hee;Kim, Young-Ki
    • Journal of Distribution Science
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    • v.12 no.11
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    • pp.55-65
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    • 2014
  • Purpose - This study aims to investigate the process of political changes in Japan, which has introduced regulatory policies for large-scale retail stores since the 1930s, as well as the examples, and suggests improvement schemes for our policies in Korea, which imposes restrictions on business hours and forced holidays in accordance with the current Distribution Industry Development Act. Research design, data, and methodology - After examining the political change processes related to large-scale retail stores in japan, this study analyzes individually regulated cases based on the ordinances enacted by each local government. Through case analysis in Japan, this study makes political suggestions that may be helpful for our country substantially. Results - Since there is an obvious possibility that our economic restrictions on business hours and mandatory holidays do not coincide with WTO GATS, it is necessary for large-scale distributors to introduce new social and environmental regulations similar to Japan, rather than imposing controls to restrict free competition and also introduce a policy to induce cooperation with small businesses for the advancement of the distribution industry. Thus, it is desirable to take measures on noise, waste, traffic, and parking for the preservation of the living environment in the surroundings when building new large-scale retail stores. It is also important to establish measures to improve the welfare of neighborhood residents and consumers, create a pleasant urban environment, and make it mandatory to make presentations at public hearings among residents. Furthermore, it should be mandatory to establish regional contribution plans when a retail store is established, and take measures to solve various civil complaints or problems that may occur after entering the market. Moreover, it is desirable for large-scale retail stores that entered the market to induce cooperation in performing various activities in the area with a strong sense that they are all members of the local economy. Conclusions - If introducing social regulations like in Japan, there is probably an advantage that the conflicts seen when large-scale retail stores enter the market are absorbed by adjusting the persons concerned within the established institution in order to establish a field to solve such conflicts systematically. In contrast, there are still concerns regarding chaotic operation without any active attempts to have a conversation with large-scale retail stores and local small merchants due to a sharp conflict among the persons concerned, and if it is a briefing session without any decision of the restrictions on their opening itself, there may be doubts with regard to their effectiveness. Moreover, if the de facto opening is restricted by the introduction of such a briefing session procedure, the choice of whether to protect the existing rights of large-scale retail stores might become problematic. However, such problems could be minimized in a way by forming a separate consultative group for all persons concerned including residents, local governments, professionals, civic organizations, small merchants, and massive retail store-related persons.

Structural and electrochemical characterization of K2NiF4 type layered perovskite as cathode for SOFCs (K2NiF4 type 층상 페롭스카이트 구조 La(Ca)2Ni(Cu)O4-δ의 SOFC 양극 특성 및 결정구조 평가)

  • Myung, Jae-ha;Hong, Youn-Woo;Lee, Mi Jai;Jeon, Dae-Woo;Lee, Young-Jin;Hwang, Jonghee;Shin, Tae Ho;Paik, Jong Hoo
    • Journal of the Korean Crystal Growth and Crystal Technology
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    • v.25 no.3
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    • pp.116-120
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    • 2015
  • $La_2NiO_{4+{\delta}}$ based oxides, a mixed electronic-ionic conductors (MIECs) with $K_2NiF_4$ type structure, have been considerably investigated in recent decades as electrode materials for advanced solid oxide fuel cells (SOFCs) due to their high electrical conductivity, and oxidation reduction reaction (ORR). In this study, structure properties of $La(Ca)_2Ni(Cu)O_{4+{\delta}}$ were studied as a potential cathode for intermediate temperature SOFCs (IT-SOFCs).

Genetic Analysis of Flower Color Traits in Calanthe discolor, C. sieboldii, and Variants Using Molecular Linkage Map (연관지도를 이용한 새우난초, 금새우난초, 변이종의 화색의 유전분석)

  • Cho, Dong-Hoon;Chung, Mi-Young;Jee, Sun-Ok;Kim, Chang-Kil;Chung, Jae-Dong;Kim, Kyung-Min
    • Journal of Life Science
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    • v.19 no.9
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    • pp.1239-1244
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    • 2009
  • This study was conducted to clarify the genetic relationship between Calanthe discolor, C. sieboldii and variants, and the cause of flower color variations by using a molecular linkage map and a quantitative trait loci (QTL) analysis for flower and lip color in Calanthe species native to Korea. Twenty plants were included in three C. discolor and three C. sieboldii, and fourteen variants were obtained from their habitat, Jeju-do in Korea. The flowers of C. discolor were brownish red, the values of Commission Internationale de I'Eclairage (CIE) Lab were between 40 and 50. The flowers of C. sieboldii were yellowish, the values of CIE Lab were between 110 and 130. The variants had various mixed colors that were thought to have originated from natural hybridization between C. discolor and C. sieboldii, and the values of CIE Lab were between 50 and 70. The colors of the lips were usually divided into white and yellow. C. discolor had a white lip, C. sieboldii had a yellow one, and the variants had a white to yellow one. The CIE Lab value of each color was 90 in white and 110 to 120 in yellow lips. A molecular linkage mapping was constructed based on the segregation of 154 RAPD markers using a MAPL program. Sixteen linkage groups containing 66 markers were established. It covered a total map distance of 220.4 cM. The distance between adjacent markers ranged from 0 to 6.6 cM, with an average distance of 3.3 cM. These markers are thought to be closely associated with flower and lip color expression. Among the 16 molecular linkage groups, 3 QTLs had flower color trait loci and 1 QTL had lip color trait loci.

Effects of the Sensory Impairment on Functioning Levels of the Elderly (노인의 감각장애와 기능상태에 관한 연구)

  • 송미순
    • Journal of Korean Academy of Nursing
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    • v.23 no.4
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    • pp.678-693
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    • 1993
  • The purposes of this study were to describe the level of vision and hearing impairments, depression and functional capacity, among Korean institutionalized elderly and to examine the relation-ship between sensory impairments, depression, and functional capacity in these people. The final pupose was to test the cognitive function path model using sensory competencies as predictors. A convenience sample of thirty nine male and 90 female subjects with a mean age of 80.5 were the subjects of this study. The subjects were tested for cognitive function, and vision and hearing impairments. Physical function and social function were measured by observation of designated task performance by the subjects. Their level of de-pression was measured using a Geriatric Depression Scale administered through an interview. Individual subjective ratings of hearing and vision were marked by the subjects, on a ladder scale. The results of the study showed that 48.8% of the subjects had a hearing impairment, 63.5% had a vision impairement, and 36.4% had both a vision and hearing impairement. The four sensory groups (no sensory impairement, hearing impairement, vision impairement, hearing and vision impairement) were tested for differences in depression, physical function, social behavior and cognitive function. The only significant difference that was found was in cognitive function, between the no sensory impairement group and the hearing and vision impairement group(F=3.25, P<.05), Subjective ratings of hearing showed a significant correlation with cognitive function(r=.34, p<.001) and with social behavior(r=.31, p<.001). There was no correlation between subjective vision ratings and cognitive function or social behavior. However there was a significant correlation between vision and hearing(r=.49, p<.001). There was also a significant negative correlation between age and vision(r=-.21, p<.01) and between age and hear-ing(r=-.34, p<.001). There was a significant correlation between depression and physical function (r=-.32, p<.001) but there was no correlation between depression and cognitive function or social behavior. Based on the literature review and the result, this study, a path model of sensory competence-> cognitive function- >social behavior was developed and tested : Perceived vision and perceived hearing were the exogenous variahles and cognitive function and social behavior were the endogeneous variables in the model. The path analysis result demonstrated an accept-able fit (GFI=.997, AGFI=.972, X$^2$=.72 (p=.396), RMSR=.019) between the data and the model. There was a significant direct effect($\beta$=.38) of perceived hearing on cognitive function. There was a significant direct effect ($\beta$=.32) of cognitive function on social behavior. The total effect of hearing on social behavior was $\beta$=.32 including the indirect effect ($\beta$=.12) . However perceived vsion had little effect ($\beta$=-.08) on cognitive function. The result of path analysis confirms that hearing levels influence cognitive function, and both hearing and cognitive function levels influence social behavior. However, vision has little effect on cognitive function or on social behavior. For the next study, a combined model of the pre viously developed environment - >depression- > physical and social function model, and the present cognitive function model, should be tested to further refine the functional capacity model. There also a need for longitudinal study of functional capacity and sencory competence in order to better understand how declining sensory competence influences functional capacity and how it effects in-creasing dependency and nursing needs in the elderly.

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Preparation and Characterization of a Sn-Anode Fabricated by Organic-Electroplating for Rechargeable Thin-Film Batteries (유기용매 전해조를 이용한 리튬이차박막전지용 Sn 음극의 제조)

  • Kim, Dong-Hun;Doh, Chil-Hoon;Lee, Jeong-Hoon;Lee, Duck-Jun;Ha, Kyeong-Hwa;Jin, Bong-Soo;Kim, Hyun-Soo;Moon, Seong-In;Hwang, Young-Ki
    • Journal of the Korean Electrochemical Society
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    • v.11 no.4
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    • pp.284-288
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    • 2008
  • Sn-thin film as high capacitive anode for thin film lithium-ion battery was prepared by organic-electrolyte electroplating using Sn(II) acetate. Electrolytic solution including $Li^+$ and $Sn^{2+}$ had 3 reduction peaks at cyclic voltammogram. Current peak at $2.0{\sim}2.5\;V$ region correspond to the electroplating of Sn on Ni substrate. This potential value is lower than 2.91 V vs. $Li^+/Li^{\circ}$, of the standard reduction potential of $Sn^{2+}$ under aqueous media. It is the result of high overpotential caused by high resistive organic electrolytic solution and low $Sn^{2+}$ concentration. Physical and electrochemical properties were evaluated using by XRD, FE-SEM, cyclic voltammogram and galvanostatic charge-discharge test. Crystallinity of electroplated Sn-anode on a Ni substrate could be increased through heat treatment at $150^{\circ}C$ for 2 h. Cyclic voltammogram shows reversible electrochemical reaction of reduction(alloying) and oxidation(de-alloying) at 0.25 V and 0.75 V, respectively. Thickness of Sn-thin film, which was calculated based on electrochemical capacity, was $7.35{\mu}m$. And reversible capacity of this cell was $400{\mu}Ah/cm^2$.