• Journal of Microbiology
• /
• 제39권3호
• /
• pp.168-176
• /
• 2001

Five different types of catalases from photosynthetic bacterium Rhodospirillum rubrum S1 grown aerobically in the dark were found in this study, and designated Catl (350 kDa), Cat2 (323 kDa), Cat3 (266 kDa), Cat4 (246 kDa), and Cat5 (238 kDa). Analysis of native PAGE revealed that Cat2, Cat3, and Cat4 were also produced in the cells anaerobically grown in the light. It is notable that only Cat2 was expressed much more strongly in response to the anaerobic condition. Enzyme activity staining demonstrated that Cat3 and Cat4 had bifunctional catalase-peroxidase activities, while Catl, Cat2, and Cat5 were typical monofunctional catalases. S1 cells grown aerobically in the presence of malate as the sole source of carbon exhibited an apparent catalase Km value of 10 mM and a Vmax of about 705 U/mg protein at late stationary growth phase. The catalase activity of Sl cells grown in the anaerobic environment exhibited a much lower Vmax of about 109 U/mg protein at late logarithmic growth phase. The catalytic activity was stable in the broad range of temperatures (30$\^{C}$-60$\^{C}$), and pH (6.0-10.0). R. rubrum S1 was much more resistant to H$_2$O$_2$in the stationary growth phase than in the exponential growth phase regardless of growth conditions. Cells of stationary growth phase treated with 15 mM H$_2$O$_2$for 1 h showed 3-fold higher catalase activities than the untreated cells. In addition, L-glutamate induced an 80-fold increase in total catalase activity of R. rubrum S1 compared with magic acid. Through fraction analyses of S1 cells, Cat2, Cat3, Cat4 and Cat5 were found in both cytoplasm and periplasm, while Catl was localized only in the cytoplasm.

• Journal of the Korean Society of Food Science and Nutrition
• /
• 제15권4호
• /
• pp.32-39
• /
• 1986

A strain of Aspergillus niger CAD 1 which produces considerable amount of beta-galactosidase was selected from extracellular beta-galctaosidase producing fungi isolated from soil. Optimal conditions for the enzyme from Aspergillus niger CAD 1 were the growth in wheat bran supplemented with 0.5% skim milk powder at $30^{\circ}C$ for 72 hrs. The crude enzyme was purified 1,387 fold through DEAE-cellulosc and Sephadex G-100 chromatographr and its recovery was 6.2%, The optimal pH and temperature for the purified enzyme were pH 4.5 ana $45^{\circ}C$, respectively. The Km and Vmax on ONPG were $3.57{\times}10^3M$ and 33.0 unit/mg protein, whereas those on lacose were $83.3{\times}10^3M$and 15.33 unit/mg protein, respectively, The activation energy for the enzyme was 9,900 cal/mol and the enzyme had no metal ion requirement for its activity and stability. The hydrolysis of lactose in skim milk, 4.8% lactose solution and acidic whey were 65%, 70% and 78% after 10 hrs incubation at $45^{\circ}C$, when 182 units of the enzyme were used 50ml of the substrate solutions.

• KSBB Journal
• /
• 제7권2호
• /
• pp.144-148
• /
• 1992

The purpose of this study is to develop a continuous process for sorbitol production using dried Zymomonas mobilis immobilized in K-carrageenan. The methods of glularaldehyde cross-linking of enzymes in CTAB (celyltrimetylammoniumbromide) treated cells immobilized in K-carrageenan showed stability for the production of sorbitol for 30 days of operation. K-carrageenan beads entrapping permeabilized cells were dried to Improve bead rigidity and storage stability. A semi-batch process with dry beads was carried out and only a small loss of enzyme activity (less than 8%) was observed during 72h. The value of Vmax for the dry K-carrageenan beads was found to be one half or that for free cells. It was shown that the productivities of the continuous process with wet K-carrageenan beads and dry beads at a dilution rate 0.1h-1 were 3.4g/L-h and 2.88h/L-h, respectively.

• Biomedical Science Letters
• /
• 제1권1호
• /
• pp.37-43
• /
• 1995

To evaluate an effect of xanthine oxidase(XO) reaction system on the carbon tetrachloride($CCl_4$) metabolism, $CCl_4$ was given twice at 0.1ml/100g body wt. at intervals of 18 hour to the rats and those pretreated with allopurinol (50mg/kg. body wt.). The influence of XO on the metabolism of $CCl_4$ was focused on the degree of liver damage and the activities of a $CCl_4$ metabolizing marker enzyme, glucose-6-phosphatase. The increasing rate of liver weight per body weight and the levels of serum alanine aminotransferase to the control group were more decreased in allopurinol-pretreated rats than in those treated with $CCl_4$ alone. The liver XO activities were more increased in $CCl_4$-treated rats than the control group and the $CCl_4$-treated rats pretreated with allopurinol showed a decreased activities of XO compared to the $CCl_4$-treated rats. The type conversion (type D --> type O) rate was more decreased tendency in allopurinol pretreated rats than those treated $CCl_4$ alone. In dialyzed liver enzyme preparations, all of the xanthine oxidase activities: $CCl_4$-treated, allopurinol and $CCl_4$-treated rats pretreated with allopurinol showed the more increased Vmax value than the control group, but similar Km value. Moreover, $CCl_4$-treated rats pretreated with allopurinol showed the more increased Vmax value than the group treated with $CCl_4$ alone. In conclusion, it can not be negate the possibility of metabolism of $CCl_4$ by the xanthine oxidase enzyme system.

• Korean Journal of Microbiology
• /
• 제34권4호
• /
• pp.225-230
• /
• 1998

This study was carried out to investigate the effect of cadmium ion on activities of cadmium-adapted malate dehydrogenase (adapted-MDH), which is defined to be an enzyme obtained from an extreme cadmium-tolerant yeast Hansenula anomaul B-7 grown in medium containing 1 mM cadmium ion. Cadmium-nonadapted malate dehydrogenase (nonadapted-MDH), which is defined to be enzyme expressed in the cells grown in $Cd^{2+}$ -free medium was also characterized by the same manner. Activities of the adapted-MDH and the nonadapted-MDH were strongly induced to 450% and to 150% in comparision with the control examined with 1 mM $Cd^{2+}$, respectively. The adapted-MDH activity was stimulated to 147%, 150%, and 135% compared with the control analyzed with 1 mM $Zn^{2+}$, 1 mM $Mn^{2+}$, and 1 mM $Ca^{2+}$, respectively and to 925%, and 250% compared with the control analyzed in the presence of 2 mM $Cd^{2+}$, and 2.5 mM $Zn^{2+}$, respectively. Km values of the adapted-MDH and the nonadapted-MDH were calculated to be the same 6.9 mM for L-malate, respectively. The Km value of the nonadapted-MDH was not changed by $Cd^{2+}$ while Vmax of the nonadapted-MDH was increased by $Cd^{2+}$. In contrast, both the Km and the Vmax values of the adapted-MDH were changed by $Cd^{2+}$.

• Journal of Environmental Health Sciences
• /
• 제16권1호
• /
• pp.67-74
• /
• 1990

An effect of carbon tetrachloride (CCl$_{4}$) was studied on the xanthine oxidase(XOD)activity of small intestine in male rats. Concomitantly a cause of increasing small intestine XOD was focused on an effect of actinomycin D and the kinetics of partial purified XOD frdm small intestine in CCl$_{4}$ intoxicated rats. An injection of CCl$_{4}$ to the rats showed an increase of small intestine XOD. In the pretreatment of actinomycin D before injection of CCl$_{4}$ to the rats, the XOD activities of small intestine were significantly decreased. In the partial purified enzyme preparation, the small intestine XOD in CCl$_{4}$ intoxicated rats showed the more increased Km and Vmax value with xanthine as substrate than that of control group.

• KSBB Journal
• /
• 제14권1호
• /
• pp.96-102
• /
• 1999

Marine bacterium Bacillus cereus ASK202 produced an extracellular agarase (E.C.3.2.1.81) which showed a high level of enzyme activity in the presence of agar and agarose. In the optimal culture conditions, the agarase production increased 7.7 folds compared with the one obtained from the basal medium. Agarase production reached upto 160 units/L after 24hr of cultivation in a modified marine medium at $25^{\circ}C$. The degree of purification increased 31.5 folds with 27.8% yield through freeze drying, DEAE Sepharose CL-6B and Superose 6HR 10/30 column chromatography. The molecular weight of the purified agarase was determined to be 90,000 daltons by gel-permeation filteration. Optimal temperature and pH for the enzyme activity were $40^{\circ}C$ and 7.8, respectively. The enzyme was stable up to $50^{\circ}C$ and at a broad pH range of 5.0-10.0. The $\beta$-agarase was activated by $Zn(NO_3)_2$, and was inhibited by $CuSO_4$ and $SnCl_2$. The Km and Vmax values of this enzyme for agarose as a substrate was $2.4mg/m\ell$ and 13.6 mg/m$\ell$, respectively.

• BMB Reports
• /
• 제33권2호
• /
• pp.172-178
• /
• 2000

The lipoxygenase was purified 35 fold to homogeneity from the Korean red potato by an ammonium sulfate precipitation and DEAE-cellulose column chromatography. The simple purification method is useful for the preparation of pure lipoxygenase. The molecular weight of the enzyme was estimated to be 38,000 by SDS-polyacrylamide gel electrophoreses and Sepharose 6B column chromatography. The purified enzyme with 2 M $(NH_4)_2SO_4$ in a potassium phosphate buffer, pH 7.0, was very stable for 5 months at $-20^{\circ}C$. Because the purified lipoxygenase is very stable, it could be useful for the screening of a lipoxygenase inhibitor. The optimal pH and temperature for lipoxygenase purified from the red potato were found to be pH 9.0. and $30^{\circ}C$, respectively. The Km and Vmax values for linoleic acid of the lipoxygenase purified from the red potato were $48\;{\mu}M$ and $0.03\;{\mu}M$ per minute per milligram of protein, respectively. The enzyme was insensitive to the metal chelating agents tested (2 mM KCN, 1 and 10mM EDTA, and 1 mM $NaN_3$), but was inhibited by several divalent cations, such as $Cu^{++}$, $Co^{++}$ and $Ni^{++}$. The essential amino acids that were involved in the catalytic mechanism of the 5-lipoxygenase from the Korean red potato were determined by chemical modification studies. The catalytic activity of lipoxygenase from the red potato was seriously reduced after treatment with a diethylpyrocarbonate (DEPC) modifying histidine residue and Woodward's reagent (WRK) modifying aspartic/glutamic acid. The inactivation reaction of DEPC (WRK) processed in the form of pseudo-first-order kinetics. The double-logarithmic plot of the observed pseudo-first-order rate constant against the modifier concentration yielded a reaction order 2, indicating that two histidine residues (carboxylic acids) were essential for the lipoxygenase activity from the red potato. The linoleic acid protected the enzyme against inactivation by DEPC(WRK), revealing that histidine and carboxylic amino acids residues were present at the substrate binding site of the enzyme molecules.

• Toxicological Research
• /
• 제13권4호
• /
• pp.371-376
• /
• 1997

To compare the severe liver damage with the slight one on the bromobeazene metabolism in rats, the animal group described as B7 group was induced the stage of slight liver damage with 7 times bromobenzene injection every other day (400 mg/Kg body wt. i.p.), whereas B40 group was induced that of more severe liver damage with bromobeazene 40 times injection as identified with determination of serum levels of alanine aminotransferase(ALT) activity and the histopathological findings. In the present experimental animal model, the decreasing rate of glutathione(GSH) and the increasing rate of glutathione S-transferase activity to the control group were higher in B7 group than B40 group. Furthermore the single dose of bromobenzene was injected to the two groups and sacrificed at 8hr and the hepatic aniline hydroxylase(AH) activity, GSH content and GST activity were determined. The increasing rate of AH activity to the control was lower in B40 group than B7 group and the decreasing rate of GSH to the control was also lower in B40 than B7 group. Moreover, B7 group showed the increased activity of hepatic GST to the control whereas B40 group showed the decrease activity of the enzyme. And Vmax value in GST was more decreased in B40 group than B7 group.

• Applied Biological Chemistry
• /
• 제36권2호
• /
• pp.105-110
• /
• 1993

Inulin fructotransferase from Enterobacter sp. S45 was purified with DEAE-cellulose column chromatography and fast protein liquid chromatography. The purified enzyme gave a single band on polyacrylamide gel electrophoresis. The molecular weight was estimated to be 42,800 by SDS-polyacrylamide gel electrophoresis. The optimal pH and temperature for the enzyme reaction were pH 5.5 and $55^{\circ}C$, respectively. $Mg^{2+}$ activated the enzyme activity, but $Fe^{3+}$, $Cu^{2+}$, $Hg^{2+}$ significantly inhibited. After exhaustive digestion of inulin by the enzyme, DFA III, sucrose, 1-kestose and nystose were produced. Sucrose, 1-kestose, raffinose and melezitose can't be used as substrates by the enzyme, but nystose and 1-F-fructofuranosyl nystose were hydrolysed. The Km and Vmax for inulin of the enzyme were 1.4 mM and $0.196\;{\mu}mole/min$, respectively.