• Title/Summary/Keyword: kinetic

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Purification and Characterization of β-N-Acetylhexosaminidase from Rice Seeds

  • Jin, Yu-Lan;Jo, Yu-Young;Kim, Kil-Yong;Shim, Jae-Han;Kim, Yong-Woong;Park, Ro-Dong
    • BMB Reports
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    • v.35 no.3
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    • pp.313-319
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    • 2002
  • N-Acetyl-$\beta$-D-hexosaminidase ($\beta$-HexNAc'ase) (EC 3.2.1.52) was purified from rice seeds (Oryza sative L. var. Dongjin) using ammonium sulfate (80%) precipitation, Sephadex G-150, CM-Sephadex, and DEAE-Sephadex chromatography, sequentially. The activities were separated into 7 fractions($F_1-F_7$) by CM-Sephadex chromatography. Among them, F6 was further purified to homogeneity with a 13.0% yield and 123.3 purification-fold. The molecular mass was estimated to be about 52 kDa on SDS-PAGE and 37.4 kDa on Sephacryl S-300 gel filtration. The enzyme catalyzed the hydrolysis of both p-nitrophenyl-N-acetyl-$\beta$-D-hexosaminide (pNP-GlcNAc) and p-nitrophenyl-N-acetyl-$\beta$-D-hexosaminide (pNP-GalNAc) as substrates, which are typical properties of $\beta$-HexNAc'ase. The ratio of the pNP-GlcNAc'ase activity to the pNP-GalNAc'ase activity was 4.0. However, it could not hydrolyze chitin, chitosan, pNP-$\beta$-glucopyranoside, or pNP-$\beta$-glucopyranoside. The enzyme showed $K_m$, $V_{max}$ and $K_{cat}$ for pNP-GlcNAc of 1.65 mM, $79.49\;mM\;min^{-1}$, and $4.79{\times}10^6\;min^{-1}$, respectively. The comparison of kinetic values for pNP-GlcNAc and pNP-GalNAc revealed that the two enzyme activities are associated with a single binding site. The purified enzyme exhibited optimum pH and temperature for pNP-GlcNAc of 5.0 and $50^{\circ}C$, respectively. The enzyme activity for pNP-GlcNAc was stable at pH 5.0-5.5 and $20-40^{\circ}C$. The enzyme activity was completely inhibited at a concentration of 0.1 mM $HgCl_$ and $AgNO_3$, suggesting that the intact thiol group is essential for activity. Chloramine T completely inhibited the activity, indicating the possible involvement of methionines in the mechanism of the enzyme.

Molecular Gene Cloning, Expression, and Characterization of Bovine Brain Glutamate Dehydrogenase

  • Kim, Dae-Won;Eum, Won-Sik;Jang, Sang-Ho;Yoon, Chang-Sik;Kim, Young-Hoon;Choi, Soo-Hyun;Choi, Hee-Soon;Kim, So-Young;Kwon, Hyeok-Yil;Kang, Jung-Hoon;Kwon, Oh-Shin;Cho, Sung-Woo;Park, Jin-Seu;Choi, Soo-Young
    • BMB Reports
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    • v.36 no.6
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    • pp.545-551
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    • 2003
  • A cDNA of bovine brain glutamate dehydrogenase (GDH) was isolated from a cDNA library by recombinant PCR. The isolated cDNA has an open-reading frame of 1677 nucleotides, which codes for 559 amino acids. The expression of the recombinant bovine brain GDH enzyme was achieved in E. coli. BL21 (DE3) by using the pET-15b expression vector containing a T7 promoter. The recombinant GDH protein was also purified and characterized. The amino acid sequence was found 90% homologous to the human GDH. The molecular mass of the expressed GDH enzyme was estimated as 50 kDa by SDS-PAGE and Western blot using monoclonal antibodies against bovine brain GDH. The kinetic parameters of the expressed recombinant GDH enzymes were quite similar to those of the purified bovine brain GDH. The $K_m$ and $V_{max}$ values for $NAD^+$ were 0.1 mM and $1.08\;{\mu}mol/min/mg$, respectively. The catalytic activities of the recombinant GDH enzymes were inhibited by ATP in a concentration-dependent manner over the range of 10 - $100\;{\mu}M$, whereas, ADP increased the enzyme activity up to 2.3-fold. These results indicate that the recombinant-expressed bovine brain GDH that is produced has biochemical properties that are very similar to those of the purified GDH enzyme.

Computations of Wave Energy by Stream Function Wave Theory (흐름함수파이론에 의한 파랑 에너지의 계산)

  • Lee, Jung Lyul;Pyun, Chong Kun
    • KSCE Journal of Civil and Environmental Engineering Research
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    • v.6 no.2
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    • pp.67-75
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    • 1986
  • This paper introduces the nonlinear Stream Function Wave Theory for design waves efficiently to compute the wave energy and energy transport quantities and to analyze the effects of nonlinearities on them. The Stream Function Wave Theory was developed by Dean for case of the observed waves with assymmetric wave profiles and of the design waves with symmetric theoretical wave profiles. Dalrymple later improved the computational procedure by adding two Lagrangian constraints so that more efficient convergence of the iterative numerical method to a specified wave height and to a zero mean free surface displacement resulted. And the Stream Function coefficients are computed numerically by the improved Marquardt algorithm developed for this study. As the result of this study the effects of nonlinearities on the wave quantities of the average potential energy density, the average kinetic energy density result in overestimation by linear wave theory compared to the Stream Function Wave Theory and increase monotonically with decreasing $L^*/L_O$ and with increasing $H/H_B$. The effects of nonlinearities on the group velocity and the wavelength quantities result in underestimation by linear wave theory and increase monotonically with increasing $H/H_B$. Finally the effect of nonlinearity on the average total energy flux results in overestimation for shallow water waves and underestimation for deep water waves by linear wave theory.

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Influence of coarse particles on the physical properties and quick undrained shear strength of fine-grained soils

  • Park, Tae-Woong;Kim, Hyeong-Joo;Tanvir, Mohammad Taimur;Lee, Jang-Baek;Moon, Sung-Gil
    • Geomechanics and Engineering
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    • v.14 no.1
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    • pp.99-105
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    • 2018
  • Soils are generally classified as fine-grained or coarse-grained depending on the percentage content of the primary constituents. In reality, soils are actually made up of mixed and composite constituents. Soils primarily classified as fine-grained, still consists of a range of coarse particles as secondary constituents in between 0% to 50%. A laboratory scale model test was conducted to investigate the influence of coarse particles on the physical (e.g., density, water content, and void ratio) and mechanical (e.g., quick undrained shear strength) properties of primarily classified fine-grained cohesive soils. Pure kaolinite clay and sand-mixed kaolinite soil (e.g., sand content: 10%, 20%, and 30%) having various water contents (60%, 65%, and 70%) were preconsolidated at different stress levels (0, 13, 17.5, 22 kPa). The quick undrained shear strength properties were determined using the conventional Static Cone Penetration Test (SCPT) method and the new Fall Cone Test (FCT) method. The corresponding void ratios and densities with respect to the quick undrained shear strength were also observed. Correlations of the physical properties and quick undrained shear strengths derived from the SCPT and FCT were also established. Comparison of results showed a significant relationship between the two methods. From the results of FCT and SCPT, there is a decreasing trend of quick undrained shear strength, strength increase ratio ($S_u/P_o$), and void ratio (e) as the sand content is increased. The quick undrained shear strength generally decreases with increased water content. For the same water content, increasing the sand content resulted to a decrease in quick undrained shear strength due to reduced adhesion, and also, resulted to an increase in density. Similarly, it is observed that the change in density is distinctively noticeable at sand content greater than 20%. However, for sand content lower than 10%, there is minimal change in density with respect to water content. In general, the results showed a decrease in quick undrained shear strength for soils with higher amounts of sand content. Therefore, as the soil adhesion is reduced, the cone penetration resistances of the FCT and SCPT reflects internal friction and density of sand in the total shear strength.

Cloning and Characterization of an Endoglucanase Gene from Actinomyces sp. Korean Native Goat 40

  • Kim, Sung Chan;Kang, Seung Ha;Choi, Eun Young;Hong, Yeon Hee;Bok, Jin Duck;Kim, Jae Yeong;Lee, Sang Suk;Choi, Yun Jaie;Choi, In Soon;Cho, Kwang Keun
    • Asian-Australasian Journal of Animal Sciences
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    • v.29 no.1
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    • pp.126-133
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    • 2016
  • A gene from Actinomyces sp. Korean native goat (KNG) 40 that encodes an endo-${\beta}$-1,4-glucanase, EG1, was cloned and expressed in Escherichia coli (E. coli) $DH5{\alpha}$. Recombinant plasmid DNA from a positive clone with a 3.2 kb insert hydrolyzing carboxyl methyl-cellulose (CMC) was designated as pDS3. The entire nucleotide sequence was determined, and an open-reading frame (ORF) was deduced. The ORF encodes a polypeptide of 684 amino acids. The recombinant EG1 produced in E. coli $DH5{\alpha}$ harboring pDS3 was purified in one step using affinity chromatography on crystalline cellulose and characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis/zymogram analysis of the purified enzyme revealed two protein bands of 57.1 and 54.1 kDa. The amino terminal sequences of these two bands matched those of the deduced ones, starting from residue 166 and 208, respectively. Putative signal sequences, a Shine.Dalgarno-type ribosomal binding site, and promoter sequences related to the consensus sequences were deduced. EG1 has a typical tripartite structure of cellulase, a catalytic domain, a serine-rich linker region, and a cellulose-binding domain. The optimal temperature for the activity of the purified enzyme was $55^{\circ}C$, but it retained over 90% of maximum activity in a broad temperature range ($40^{\circ}C$ to $60^{\circ}C$). The optimal pH for the enzyme activity was 6.0. Kinetic parameters, $K_m$ and $V_{max}$ of rEG1 were 0.39% CMC and 143 U/mg, respectively.

Studies on the Ethanol Production by Clostridium thermosaccharolyticum (Clostridium thermosaccharolyticum에 의한 에탄올생산에 관한 연구)

  • 조은경;이윤광;변유량;유주현
    • Microbiology and Biotechnology Letters
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    • v.13 no.4
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    • pp.397-402
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    • 1985
  • The fermentation of various sugars by C. thermosaccharolyticum was examined under pH controlled, anaerobic condition. The kinetic model for Product formation at various sugars was the combination of growth and non-growth associated mode. In the utilization of a single sugar, glucose was the best carbon source for growth. The specific growth rate of glucose, xylose and cellobiose were 0.363 h$^{-1}$, 0.242 h$^{-1}$ and 0.144 h$^{-1}$ respectively. The production of ethanol from glucose showed a negatively growth associated mode, so the higher growth rate decreased the productivity of ethanol. The maximum concentrations of the produced ethanol were 2.42 g/l, 3.76 g/l, and 3.4 g/l on glucose, xylose, and cellobiose. No glucose was detected during cellobiose fermentation. Sequential utilization of sugars was observed in the mixtures of glucose, xylose and cellobiose. It preferred glucose, followed by xylose and then cellobiose. The presence of other sugars had little or no effect on the rate of another sugar utilization. Cell lysis at the end of fermentation occured more slowly in the mixtures of sugars than a single sugar.

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In vitro Evaluation of Phalaris minor Seeds as Livestock Feed

  • Kaur, J.;Pannu, M.S.;Kaushal, S.;Wadhwa, M.;Bakshi, M.P.S.
    • Asian-Australasian Journal of Animal Sciences
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    • v.19 no.3
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    • pp.363-367
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    • 2006
  • The nutritional worth of Phalaris minor seeds was assessed in comparison to conventional cereal grains like maize and wheat. P. minor seeds had higher total ash and cell wall constituents as compared to wheat and maize grains. The CP content of P. minor was comparable to wheat grains but higher than maize grains. The in vitro studies revealed that the net gas production and availability of ME from P. minor was comparable to that of maize but the digestibility of nutrients was significantly (p<0.05) lower than that of conventional cereal grains. The digestion kinetic parameters for DM and CP revealed that P. minor had the highest (p<0.05) soluble fraction (a) followed by wheat and maize. Reverse trend was observed for insoluble but potentially degradable fraction (b). The effective and true DM and CP degradability was significantly (p<0.05) higher in wheat grains followed by that in P. minor and maize grains. The digestibility of OM and NDF was not affected by replacing cereal grains in concentrate mixture with P. minor seeds up to 75 per cent level. But the availability of ME from concentrate mixtures was comparable to control only up to 50% level of replacement. Replacement of cereal grains with P. minor did not affect the rapidly soluble fraction and insoluble but potentially degradable fraction of concentrate mixture containing P. minor up to 75 per cent, but it was depressed significantly at 100% replacement level. The effective and true degradability of DM of concentrate mixtures containing P. minor from 50 to 100 per cent was comparable to that of conventional concentrate mixture (CCM). The wheat based concentrate mixtures showed higher net gas production (208 vs. 201 ml/g DM/24 h), digestibility of nutrients and ME availability (9.64 vs. 9.54 MJ/kg DM) as compared to maize based concentrate mixture. The wheat based concentrate mixture had significantly (p<0.05) higher rumen undegradable fraction and effective degradability. The data conclusively revealed that conventional cereal grains could be replaced with P.minor seeds up to 75 per cent without affecting the availability of nutrients.

Effects of low NaNO2 and NaCl concentrations on Listeria monocytogenes growth in emulsion-type sausage

  • Lee, Jeeyeon;Gwak, Eunji;Lee, Heeyoung;Ha, Jimyeong;Lee, Soomin;Kim, Sejeong;Oh, Mi-Hwa;Park, Beom-Young;Choi, Kyoung-Hee;Yoon, Yohan
    • Asian-Australasian Journal of Animal Sciences
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    • v.30 no.3
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    • pp.432-438
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    • 2017
  • Objective: The objective of this study was to evaluate the effect of combinations of $NaNO_2$ and NaCl concentrations on Listeria monocytogenes (L. monocytogenes) growth in emulsion-type sausage. Methods: Emulsion-type sausages formulated with different combinations of $NaNO_2$ (0 and 10 ppm) and NaCl (1.00%, 1.25%, and 1.50%) were inoculated with a five-strain L. monocytogenes mixture, and stored at $4^{\circ}C$, $10^{\circ}C$, and $15^{\circ}C$, under aerobic or vacuum conditions. L. monocytogenes cell counts were measured at appropriate intervals, and kinetic parameters such as growth rate and lag phase duration (LPD) were calculated using the modified Gompertz model. Results: Growth rates increased (0.004 to 0.079 Log colony-forming unit [CFU]/g/h) as storage temperature increased, but LPD decreased (445.11 to 8.35 h) as storage temperature and NaCl concentration increased. The effect of combinations of NaCl and low-$NaNO_2$ on L. monocytogenes growth was not observed at $4^{\circ}C$ and $10^{\circ}C$, but it was observed at $15^{\circ}C$, regardless of atmospheric conditions. Conclusion: These results indicate that low concentrations of $NaNO_2$ and NaCl in emulsion-type sausage may not be sufficient to prevent L. monocytogenes growth, regardless of whether they are vacuum-packaged and stored at low temperatures. Therefore, additional techniques are necessary for L. monocytogenes control in the product.

Frictional resistance of different ceramic brackets and their relationship to the second order angulation between bracket slot and wire (세라믹 브라켓의 종류 및 브라켓 슬롯과 와이어 각도에 따른 마찰 저항 차이)

  • Choi, Yoon-Jeong;Park, Young-Chel
    • The korean journal of orthodontics
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    • v.36 no.3 s.116
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    • pp.207-217
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    • 2006
  • Although ceramic brackets have been used widely for improved esthetics during treatment, ceramic brackets have some inherent problems; brittleness, attrition of the opposing teeth and high frictional resistance. This study was performed to understand the frictional resistance of the ceramic brackets, as well as to be a helpful reference for finding the solutions to the problem of frictional resistance. Three different kinds of brackets were used; metal bracket, polycrystalline ceramic brackets with a metal slot to reduce the high frictional resistance and monocrystalline ceramic brackets. The brackets were tested with a $.019{\times}.025$ stainless steel wire with a second order angulation of $0^{\circ}\;and\;10^{\circ}$, and the static and kinetic frictional forces were measured on the universal testing machine. The results of this study showed that the ceramic brackets, especially the monocrystalline ceramic bracket without a metal slot, generated higher frictional resistance than the metal bracket, and the frictional resistance was increased as the angulation between the bracket slot and the wire increased. Therefore, the development of the ceramic bracket with reduced frictional resistance and the prevention of excessive crown tipping during orthodontic treatment will lead to the simultaneous attainment of more efficient and improved esthetic treatment goals.

Wavelength-resolved Thermoluminescence of Chemical-vapor-deposited Diamond Thin Film (화학증착된 다이어몬드 박막의 파장 분해된 열자극발광)

  • Cho, Jung-Gil;Yi, Byong-Yong;Kim, Tae-Kyu
    • Progress in Medical Physics
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    • v.12 no.1
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    • pp.1-8
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    • 2001
  • Diamond thin films were synthesized by a chemical vapor deposition (CVD). Raman spectrum showed the diamond line at 1332 $cm^{-1}$ / and x-ray diffraction pattern exhibited a strong (111) peak of diamond. The scanning electron microscopy analysis showed that the CVD diamond thin film was grown to be unepitaxial crystallites with pyramidal hillocks. A wavelength-resolved thermoluminescence (TL) of the CVD diamond thin film irradiated with X-rays showed one peak at 430 nm around 560 K. The glow curve of the CVD diamond thin film produced one dominant 560-K peak that was caused by first-order kinetics. Its activation energy and the escape frequency were calculated to be 0.92 ~ 1.05 eV and 1.34 $\times$ 10$^{7}$ sec$^{-1}$ , respectively. The emission spectrum at 560 K was split into 1.63-eV, 2.60-eV, and 3.07-eV emission bands which is known to be attribute to silicon-vacancy center, A center, and H3 center, respectively.

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