• Title/Summary/Keyword: Lineweaver-Burk plot

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Mixed-type Inhibition of Human Hepatic Cytochrome P450 1-Catalyzed Ethoxyresorufin O-deethylation by Volatile Allyl Sulfides

  • Kim, Hyun-Jung;Chun, Hyang-Sook
    • Food Science and Biotechnology
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    • v.14 no.2
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    • pp.297-300
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    • 2005
  • Effects of allyl sulfides on kinetic behavior of cytochrome P450 1 (CYP1)-catalyzed ethoxyresorufin O-deethylase (EROD) activity were studied using microsomes from benzo[a]pyrene-treated human hepatoma cells. Apparent $K_m$ and $V_{max}$ values were calculated as $2.8\;{\mu}M$ and $3.0\;{\mu}mol$ resorufin/min/mg protein based on Lineweaver-Burk plot of microsomal EROD activity, respectively. Diallyl disulfide (DADS) and diallyl trisulfide (DATS) affected $K_m$ and $V_{max}$ values of EROD activity and acted as mixed-type inhibitors for CYP1 isozymes. Apparent Ki values of DADS and DATS were calculated as 1.07 and 0.88 mM, respectively, by re-plotting slopes of Lineweaver-Burk plot and inhibitor concentrations.

Kinetics on the Specificity of Enzymatic Hydrolysis of Chitin (Chitin의 효소적 가수분해 특성에 대한 속도론적 연구)

  • Lee, Eun-Young;Kim, Kwang
    • KSBB Journal
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    • v.13 no.1
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    • pp.44-51
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    • 1998
  • Hydrolysis and adsorption reversibility experiments were run for initial enzyme activity of 4.48, 9.65, 11.19 and 17.14U/mL at a temperature 30$^\circ C$. The chitin particle size corresponded to a mean particle diameter of 0.127mm, and the initial concentration of chitin was 10mg/mL. After approximately 2hrs, the enzyme activity remained constant in a speudo-steady state. The amounts in the bulk [E] and the amounts of enzyme adsorbed on the chitin surface [E] are plotted on Lineweaver-Burk plot to yield a linear relationship with a correlation coefficient of 0.99, a slope of 2.79cm$^-1$ and an intercept of 0.08$\textrm{cm}^2$/U. From this parameters, the values of [E$_T$] and $K_E$ were calculated to be 12.5U/cm$^2$ and 34.88U/mL. respectively, Adsorption isotherm of the enzyme on the particles showed a well developed plateau of 1.35$\times$10$^-3$, 4.72$\times$10$^-3$, 4.42$\times$10$^-3$, 8.58$\times$10$^-3$U/cm$^2$ at 30$^\circ C$. To determine the specificity of chitinase for crystalline chitin, the free energy of adsorption was measured, and its was determined as about -14.62~-18.8kJ/mol.

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Purification and Characterization of Cholesterol Oxidase Produced by Streptomyces sp. No.4 (방선균 Streptomyces sp. No.4가 생산하는 Cholesterol Oxidase의 정제 및 특성)

  • 김현수;고희선
    • KSBB Journal
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    • v.14 no.3
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    • pp.322-327
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    • 1999
  • The cholesterol oxidase(EC.1.1.3.6) produced from Streptomyces sp. No.4 which isolated from soil was purified and investigated for the enzymatic properties. The enzyme was purified specifically by cholesterol affinity column chromatography with a yield of 28.3%. The purified enzyme showed a single polypeptide on SDS-PAGE and the molecular weight was estimated to be 60,000 daltons. The enzyme activity was strongly inhibited by metal ions such as $HgCl_2$ and $CuSO_4$. Dithiothreitol and mercaptoethanol inhibited the enzyme activity at concentration of 1mM. The Michaelis constant(Km) for cholesterol was found to be 1.38mM by Lineweaver-Burk plot analysis. Amino acid analysis showed that the enzyme protein was composed of 416 amino acid residues including 52moles of glycine and 19moles of tryptophane.

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Isolation and Characterization of α-Glucosidase Inhibitor Produced by Bacillus sp. SKU31-1 Strain (Bacillus sp. SKU31-1가 생산하는 α-Glucosidase 저해제 분리 및 특성 조사)

  • Kim, Shin-Duk
    • Korean Journal of Microbiology
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    • v.50 no.4
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    • pp.381-383
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    • 2014
  • In the course of screening for ${\alpha}$-glucosidase inhibitor produced by microorganism, the active compound was isolated from the culture filtrate of Bacillus sp. SKU31-1 using a series of chromatography procedures. The structure of the active compound was elucidated as 5-amino-1-hydroxymethyl-1, 2, 3, 4-cyclohexanetetrol on the basis of spectroscopic evidence obtained and comparison with data from the literature. The active compound showed potent inhibitory activity against ${\alpha}$-glucosidase with an $IC_{50}$ value of $1.9{\mu}M$ for maltose and 4.9 mM for sucrose. A Lineweaver-Burk plot indicated that its inhibition of ${\alpha}$-glucosidase was competitive, with a $K_i$ value of 0.15 mM.

Electrochemical Properties of HRP Immobilized Biosensor bound with EPDM (EPDM으로 결합된 HRP 고정 바이오센서의 전기화학적 특성)

  • Yoon, Kil-Joong
    • Elastomers and Composites
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    • v.42 no.2
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    • pp.112-118
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    • 2007
  • An HRP immobilized carbon paste electrode, which was bound by EPDM, was newly fabricated and its electrochemical properties were investigated for the purpose of validating the new possibility for the practical use of biosensor. In the experimental range of substrate concentration, Lineweaver-Burk plot of the signal showed a good linearity. This means that HRP was embedded effectively to preserve its identity in the bulk of composite electrode materials and EPDM is a recommendatory binder. When the electrode was run at low operating potential($0.0\sim-1.0$ V vs. Ag/AgCl), it showed a high sensitivity and a good reproducibility. Especially the mechanical stability of the dried rubber was a remarkable breakthrough to get over a difficulty in putting the carbon-paste electrode bound with silicon oil to real use.

Electrochemical properties of the mugwort-embedded biosensor for the determination of hydrogen peroxide (쑥을 이용한 과산화수소 정량 바이오센서의 전기화학적 성질)

  • Lee, Beom-Gyu;Park, Sung-Woo;Yoon, Kil-Joong
    • Analytical Science and Technology
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    • v.19 no.1
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    • pp.58-64
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    • 2006
  • A mugwort-tissue-based modified carbon paste electrode was constructed for the amperometric detection of hydrogen peroxide and its electrochemical properties are described. Especially the amperometric signal was very stable and bigger than any other enzyme electrode studied in this lab. The effect of tissue composition on the response was linear within the wide range of experiment and the linearity of Lineweaver-Burk plot showed that the sensing process of the biosensor is by enzymatic catalysis. And pH dependent current profile connoted that two isozymes are active in this system.

Inhibitory Effect of Buthus martensi Karsch Extracts on ${\alpha}$-Glucosidase Enzyme

  • Kim, Eun-Ok;Kim, Shin-Duk
    • International Journal of Industrial Entomology and Biomaterials
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    • v.15 no.2
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    • pp.161-164
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    • 2007
  • While searching for ${\alpha}$-glucosidase inhibitors, the active compound was found in a methanol extract of Burthus martensi Kirsch. The separation of the active compound was performed using various chromatography methods and the physico-chemical properties of the purified compound were characterized. The compound showed very potent inhibitory activity against ${\alpha}-glucosidase$ with an $IC_{50}$ value of $5.3\;{\mu}g/ml$. Lineweaver-Burk plot indicated that its inhibition of ${\alpha}-glucosidase$ was competitive.

Characteristics of Lipoxygenase in Black Rice (진도산 흑미의 lipoxygenase의 특성)

  • 이유석;송선주;이종욱
    • Food Science and Preservation
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    • v.6 no.2
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    • pp.216-220
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    • 1999
  • Lipoxygenase(LOX) activity of black rice(Chindo) was measured by spectrophotometric method at In m. Studies at different pH levels revealed that the optimal activity was exhibited at pH 7.0 with 24.97 unit/mg. Enzyme activity was tested at different concentration of the substrate. The apparent Vmax and Km values were determined from the Lineweaver-Burk plot to be 53.85 unit/mg and 0.21 mM. Enzyme activity due to storage temperature (-40, 4 and 25$^{\circ}C$) and period were decreased at all storage temperature. LOX activity of black rice was significantly decreased during the microwave heating.

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Electrochemical Properties of Biosensor with Butyl Rubber Binder (부틸고무를 결합재로 사용한 바이오센서의 전기화학적 성질)

  • Choi, Sei-Young;Yoon, Kil-Joong
    • Elastomers and Composites
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    • v.41 no.4
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    • pp.231-237
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    • 2006
  • An enzyme electrode bound by butyl rubber was newly constructed for the determination of hydrogen peroxide and for the practical application as a biosensor. Then its electrochemical properties were investigated. It produced a hundreds-fold increased signal compared to the plant or animal tissue based biosensor studied previously and could be run at between $0.0{\sim}-1.00\;V$(vs. Ag/AgCl). The relationship between signal and electrode potential was linear in the experimental range of potential. It showed a detection limit of $3.0{\times}10^{-4}\;M$ and a very good linearity of Lineweaver-Burk plot giving the proof of a good enzyme immobilization. Especially, both the reproducibility of signal current due to its high sensitivity and mechanical stability presented a new possibility for the practical use of biosensor bound with butyl rubber.

Conversion of water-insoluble components of the basidiocarps of ganoderma lucidum to water-soluble components by hydrolyzing with chitinase

  • Cheong, Jae-Yeon;Park, Won-Bong
    • Archives of Pharmacal Research
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    • v.19 no.4
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    • pp.326-334
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    • 1996
  • We investigated the optimum conditions for conversion of water-insoluble components of basidiocarps of Ganoderma lucidum to water-soluble components by hydrolyzing with chitinase. We also tried it with Ganoderma luciclum residue remaining after extracting hot water-soluble components of Ganoderma lucidum. After hydrolyzing under optimum conditions (20 ppm chitinase, 2% Ganoderma lucidum or 6% Ganoderma lucidum residue, at pH 3 and at $ 35^{\circ}C$), the contents of total water-soluble components (polysaccharide or protein) were measured, and it was found that the contents of water-soluble components increased to 1.5-2.7 fold. Michaelis constant, $K_m$ and maximum rate, $V_max$ calculated by Lineweaver-Burk plot for hydrolysis of Ganoderma lucidum were 1.75% and 0.02%/min respectively and those for hydrolysis of Ganoderma lucidum residue were 53.15% and 0.53%/min respectively The protein-bound polysaccharide was isolated after hydrolysis and molecular weights were measured by Sepharose CL-4B gel filtration and compared with the molecular weights of polysaccharide before hydrolysis.

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