• Korean Journal of Microbiology
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• v.18 no.1
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• pp.7-14
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• 1980

A thermostrable ${\beta}-galactosidase$ (${\beta}-galactoside$ galactohydorlase, EC 3.2.1.23) was inducible in Bacillus coagulans by lactose and D-glactose. The enzyme was purified 87 fold, and the optimum temeprature and pH for actiivity were determined to be $60^{\circ}C$ and pH 7.5, respectively. Kinetic determinations at $55^{\circ}C$ established a Km of 3.3mM for the chromogenic substrate onitorphenyl ${\beta}-D-galactopyranoside$ (ONPG). Galactose and lactose were competitive inhibitors with Ki of 6.1mM and 4.9mM, respectively. The enzyme ws relatively thermostable. The crude enzyme was inactivated about 20% after 20 min of exposure at $60^{\circ}C$ and the purified was about 50%. Maximal enzyme activity required $Mn^{++}$, and for the thermal stabilization $Fe^{++}\;and\;Ca^{++}$ were necessary.

• Bulletin of the Korean Chemical Society
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• v.23 no.8
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• pp.1057-1061
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• 2002

The Hafnia alvei aspartase activity with acetic anhydride treatment gradually increased and reached 7.5-fold that of the native one. The activity of the acetylated aspartase was a little higher than that of the native enzyme, indicating that the cooperativity between a substrate and enzyme is increased. The optimum temperature of the native asparatse was $45^{\circ}C$, and that of the acetylated enzyme shifted to $40^{\circ}C.$ The pH vs. the activity profile of the acetylated asparatse was also different from that of the native enzyme. The initial velocity pattern of the acetylated aspartase intersects to the left of the ordinate, indicating the sequential kinetic mechanism other than a rapid equilibrium ordered one. The reciprocal plots for aspartate of the native aspartase were curved, but those of the acetylated aspartase were linear, indicating the Michaelis-Menten kinetics. The helical content of the acetylated aspartase was rather decreased to $9{\textperthousand}$ than that $(63{\textperthousand})$ of the native one.

• Proceedings of the Korean Institute of Electrical and Electronic Material Engineers Conference
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• 2000.07a
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• pp.909-912
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• 2000

We synthesized polypyrrole (PPy) nanotubules by oxidative polymerization of the pyrrole monomer within the pores of a polycarbonate template. The electrochemical behavior was investigated using cyclic voltammetry. The redox potential was about -0.5 V vs. Ag/AgCl reference electrode, while the potential was about 0 V for PPy film. It is considered as the backbone grows according to the pore wall. Therefore, it is possible to be arranged regularly. That leads to improvement in the electron hopping. By electrochemical doping of glucose oxidase (GOx) on PPy nanotubules, an enzyme electrode has been fabricated. The kinetic parameter of biochemical reaction with glucose was evaluated. The formal Michaelis constant and maximum current calculated by computer were about 11.4 mmol $dm^3$ and 170.85 A respectively. Obviously, an affinity for the substrate and current response of the PPy nanotubules enzyme electrode are rather good, comparing with that of PPy film.

• Bulletin of the Korean Chemical Society
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• v.3 no.3
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• pp.122-129
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• 1982

In order to extend our understanding on the multiple inhibition enzyme kinetics, a general equation of an enzyme reaction in the presence of two different reversible inhibitors was derived by what we call "match-box mechanism" under the combined assumption of steady-state and quasi-equilibrium for inhibitor binding. Graphical methods were proposed to analyze the multiple inhibition of an enzyme by any given sets of different inhibitors, i.e., competitive, noncompetitive, and uncompetitive inhibitors. This method not only gives an interaction factor $({\alpha})$ between two inhibitors, but also discerns ${\alpha}_1$ and ${\alpha}_2$ with and without substrate binding, respectively. The factors involved in the dissociation constants of inhibitors can also be evaluated by the present plot. It is also shown that the present kinetic approach can be extended to other forms of activators or hydrogen ions with some modification.

• Archives of Pharmacal Research
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• v.5 no.2
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• pp.45-52
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• 1982

Staibilization effect of Panax ginseng C. A. Meyer on .betha.-D-Galactosidase inactivation was proved by kinetic studies of thermal inactivation of the enzyme. The water extract Panax ginseng C. A. Meyer showed stabilization activity at minimal concentration of 10ppm. The methanolic extract was purified to obtain ginseng saponins, and two groups of the ginsenosides, i. e. protopanaxadiol and protopanaxatriol were isolated. They also showed a protective effect against the thermal and chemical inactivation of the enzyme; p-chloromercuribenzoic acid and hydroxylamine known as protein modifier greatly inactivated the enzyme but inactivation was significantly balocked by the ginseng component MG$^{2+}$, known as a cofactor, stabilized the enzyme and the poor stabilization effect by it was potentiated by ginseng components.s.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 1978.10a
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• pp.206.4-206
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• 1978

By utilizillg whole cell enzyme of the Xantho-monas citri IFO 3835, cephalexin is synthesized directly from 7-amino-deacetoxy cephalosporanic acid (7-ADCA) and phenyl glycine methyl ester (PGM). To date, cephalexin has been manufactu-red by chemical process involving fairly large number of steps to protect the amino group of phenly glycine and carboxyl group of 7-ADCA. However, the enzymatic process involves only a single step with 85％ conversion in 90 minutes. The fermentation variables studied indicate that oxygen transfer is limiting step in the enzyme production. Optimum conditions for enzymatic reaction were 37 C, pH 6.0, and the optimum substrate molar ratio of PGM to 7-ADCA was 2. Other variables that are related to the biochemical properties of whole cell enzyme temperature stability, pH stability, kinetic constants, reusing effect, enzyme loading effect were also evaluated.

• Journal of Life Science
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• v.23 no.12
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• pp.1471-1476
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• 2013

The present study deals with the immobilization of Kluyveromyces lactis ${\beta}$-galactosidase on a weak ionic exchange resin (Duolite A568) as polymer support. ${\beta}$-Galactosidase was immobilized using the adsorption method. A kinetic study of the immobilized enzyme was performed in a packed-bed reactor. The adsorption of the enzyme followed a typical Freundlich adsorption isotherm. The adsorption parameters of k and n were 14.6 and 1.74, respectively. The initial rates method was used to characterize the kinetic parameters of the free and immobilized enzymes. The Michaelis-Menten constant ($K_m$) for the immobilized enzyme (120 mM) was higher than it was for the free enzyme (79 mM). The effect of competitive inhibition kinetics was studied by changing the concentration of galactose in a recycling packed-bed reactor. The kinetic model with competitive inhibition by galactose was best fitted to the experimental results with $V_m$, $K_m$, and $K_I$ values of 46.3 $mmolmin^{-1}mg^{-1}$, 120 mM, and 24.4 mM, respectively. In a continuous packed-bed reactor, increasing the flow rate of the lactose solution decreased the conversion efficiency of lactose at different input lactose concentrations. Continuous operation of 11 days was conducted to investigate the stability of a long-term operation. The retained activity of the immobilized enzymes was 63% and the half-life of the immobilized enzyme was found to be 15 days.

• BMB Reports
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• v.41 no.11
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• pp.790-795
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• 2008

An inducible lysine 6-dehydrogenase (Lys 6-DH), which catalyzes the oxidative deamination of the 6-amino group of L-lysine in the presence of $NAD^+$, was purified to homogeneity from Achromobacter denitrificans, yielding a homodimeric protein of 80 kDa. The enzyme was specific for the substrate L-lysine and $NAD^+$ served as a cofactor. The dimeric enzyme associated into a hexamer in the presence of 10 mM L-lysine. The $K_m$ values for L-lysine and $NAD^+$ were 5.0 and 0.09 mM, respectively. The lys 6-dh gene was cloned and overexpressed in E. coli. The open reading frame was 1,107 nucleotides long and encoded a peptide containing 368 amino acids with 39,355 Da. The recombinant enzyme was purified to homogeneity and characterized. Enzyme activities and kinetic properties of the recombinant enzyme were almost the same as those of the endogenous enzyme obtained from A. denitrificans. Crystals of the enzyme were obtained using the hanging drop method.

• Korean Journal of Ecology and Environment
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• v.33 no.4 s.92
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• pp.380-386
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• 2000

Alkaline phosphatase ($AP_{ase}$) activity was studied for the pure cultures and natural communities of phytoplankton. The Michaelis-Menten constant ($K_m$) showed large variation with species. Some green algae showed large $K_m$ ($650\;{\mu}M$ for Selenastrum capriconutum). Chlorella sp. and Nitzschia palea showed smaller Km (respectively1.7, 2.0 HM) than those of other species examined. The extracellular free enzyme in the filtrate of Anabaena flos-aquae showed smaller Km ($52\;{\mu}M$) than that of cellbound form ($276\;{\mu}M$). The $K_m$ ($12.0\;{\mu}M$) of summer phytoplankton in Lake Soyang. when Anabaena sp. was dominant species, was larger than that (1.5 HM) of spring phytoplankton when Asterionella sp. was the dominant. Although maximum activity($V_{max}$) in Lake Soyang was affected by the concentration of DIP within the lake, but the $V_{max}$ always varied not with the DIP concentration of the lake. Induction of $AP_{ase}$ may be more affected by the phosphate content within the cell of phytoplankton than by the concentration of DIP within the lake, The extracellular free $AP_{ase}$ activity accounted for $36{\sim}97%$ of total activity from fall to spring turnover in Lake Soyang. The $K_m$ ($1.1{\mu}3.5\;M$) of extracelluar free enzyme were simillar to those ($0.7{\mu}3.5\;M$) of the total activity. This indicates that the extracelluar free enzyme was derived from phytoplankton.

• Archives of Pharmacal Research
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• v.3 no.1
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• pp.7-12
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• 1980

Ampliciline was synthesized from 6-amino-pencillanic acid (6-APA) and D-.alpha. phenylglycine methyl ester by using amplicilin synthesizing enzyme from Peudomonas melanogenum (IAM 1655). The whole cell enzyme was immobilized by entrapping it in the polyacrylamide gel lattices. The polymer used in the enzyme entrapment was made from 150 mg per ml of acrylamide monomer and 8 mg per ml of N, N'-methylenebisacrylamide. About 200 mg/whole cell enzyme was mixed in the polymer for entrapment. The maximal activity retention after immobilization was 56%. The optimal pH values for the whole cell enzyme and the immobilized whole cell enzyme were 6.0 and 5.9, respectively. The optimal temperature for the enzyme activity were the same for both type of preparations. The enzyme stabilities against pH and heat increased for immobilized whole cell enzyme. Immobilized cell was more stable especially in the acidic condition while both type were found to be very suceptible to thermal inactivation at a temperature above 4.deg.C. The kinetic constants obtained from Lineweaver-Burk plot based on two substate reaction mechanism showed somewhat higher value for immobilized whole cell enzyme as compared to the whole cell enzyme : the Km value for 6-APA were 7.0 mM and 12.5 mM while Km values for phenylglycine methyl ester were 4.5 mM and 8.2 mM, respectively. Using the immobilized whole cell enzyme packed in a column reactor, the productivity of ampiciline was studied by varying the flow rate of substrate solution. At the space velocity, SV, 0.14 hr$^{-1}$ the conversion was 45%. Operational stability found in terms of half life was 30 hr at SV = 0.2 hr.