• BMB Reports
• /
• v.44 no.1
• /
• pp.64-69
• /
• 2011

A peroxidase (PD-cP; 0.47 mg/100 g leaves) was purified from autumn leaves of Phytolacca dioica L. and characterized. PD-cP was obtained by acid precipitation followed by gel-filtration and cation exchange chromatography. Amino acid composition and N-terminal sequence of PD-cP up to residue 15 were similar to that of Spinacia oleracea (N-terminal pairwise comparison showing four amino acid differences). PD-cP showed a molecular mass of approx. 36 kDa by SDS-PAGE, pH and temperature optima at 3.0 and $50.0^{\circ}C$, respectively and seasonal variation. The Michaelis-Menten constant ($K_M$) for $H_2O_2$ was 5.27 mM, and the velocity maximum ($V_{max}$) $1.31\;nmol\;min^{-1}$, while the enzyme turnover was $0.148\;s^{-1}$. Finally, the presence of $Ca^{2+}$ and $Mg^{2+}$ enhanced the PD-cP activity, with $Mg^{2+}$ 1.4-fold more effective than $Ca^{2+}$.

• Current Research on Agriculture and Life Sciences
• /
• v.31 no.4
• /
• pp.227-232
• /
• 2013

We studied antioxidant activity and inhibitory effect of ${\alpha}$-glucosidase from aqueous, ethanolic and methanolic fractions of Galla rhois. In FRAP and ORAC assay for measuring antioxidant activity, we confirmed that Galla rhois extracts had strong antioxidant activity and ethanolic and methanolic extracts were relatively stronger than aqueous extract. We used trolox as a positive control. In order to measure the inhibitory effect of ${\alpha}$-glucosidase, we compared acarbose and Galla rhois extracts. As a result of ${\alpha}$-glucosidase inhibitory assay, aqueous, ethanolic and methanolic extracts of Galla rhois showed high inhibitory activitity and ethanolic and methanolic extracts were relatively stronger than aqueous extract. The 50% inhibitory concentrations (IC50s) of acarbose, aqueous, ethanolic and methanolic fractions were 0.45 mM, $0.53{\mu}g/ml$, $0.415{\mu}g/ml$ and $0.37{\mu}g/ml$, respectively. These results suggest that Galla rhois extracts can be a clinically useful anti-diabetic ingredient, indicating that it needs to be fractionated and isolated and should be further investigated.

• Journal of the Korean Society of Clothing and Textiles
• /
• v.20 no.3
• /
• pp.550-559
• /
• 1996

The Effect of protease (subtilisin Carlsberg) on the removal of hemoglobin as protein soil was studied. The hydrolysis characteristics of subtilisin Carlsberg was examined by electrophoretic techniques. The fragmentation patterns of hemoglobin were analyzed by SDS-PAGE. The hydrolysis efficiency was evaluated by analysis of protein bands shown on gels before and after hydrolysis by using densitometer. 1. The hydrolysis of hemoglobin by subtilisin Carlsberg was increased markedly with the increase of the enzyme concentration. 2. The hydrolysis of hemoglobin by subtilisin Carlsberg was effectively increased in proportion to increasing of the hemoglobin concentration up to a certain point, but it began to decrease above the point. 3. The hydrolysis of hemoglobin by subtilisin Carlsberg followed the first order kinetics, yielding a rate constant of $4.05\time10^{-4}S^{-1}s$. 4. The hydrolysis of hemoglobin by subtilisin Carlsberg was highest at $50^{\circ}C$ and was decreased markedly at $80^{\circ}C$. 5. The hydrolysis of hemoglobin was comparatively low at pH 7.0~8.0, and highest at pH 11.0.

• Bulletin of the Korean Chemical Society
• /
• v.35 no.2
• /
• pp.425-430
• /
• 2014

Amyloidogenic proteins often exhibit fibrillar polymorphism through alternative assembly processes, which has been considered to have possible pathological implications. Here, firefly luciferase (LUC) is shown to induce amyloid polymorphism of ${\alpha}$-synuclein, the major constituent of Lewy bodies found in Parkinson's disease, by acting as a novel template. The drastically accelerated fibrillation kinetics of ${\alpha}$-synuclein with LUC required the nucleation center produced by the active enzyme of LUC. Fluorescent dye binding, transmission electron microscopy, and Fourier transformed infrared spectroscopy revealed the morphologically distinctive amyloid fibrils of ${\alpha}$-synuclein prepared in the absence or presence of LUC. As the altered morphological characteristics became inherent to the mature fibrils, those properties were inherited to next-generations via nucleation-dependent fibrillation process. The seed control, therefore, would be an effective means to modify amyloid fibrils with different biochemical characteristics. In addition, the LUC-directed amyloid fibrillar polymorphism also suggests that other cellular biomolecules including enzymes in general are able to diversify amyloid fibrils, which could be self-propagated with diversified biological activities, if any, inside cells.

• Journal of Microbiology
• /
• v.35 no.1
• /
• pp.25-29
• /
• 1997

Microbial reduction of hexavalent (VI) to trivalent (III) chromium decreases its toxicity by two orders of magnitude. In order to investigate the nature of Cr-reduction, Cr-resistant Pseudomonas aeruginosa HP014 was isolated and tested for its reduction capability. At the concentration of 0.5 mM Cr(VI), cell growth was not inhibited by the presence of Cr(VI) in a liquid medium, and Cr(VI) reduction was accompanied by ell growth. When cell-free extract was tested, the reduction of Cr(VI) showed a saturation kinetics with the maximum specific activity of 0.33 .mu.mol min$\^$-1/ mg$\^$-1/ cell protein, and an apparent K. of 1.73 mM Cr(VI). The activity required either NADH or NADPH as an electron donor. However, NADPH gave 50% as mush activity as sequently the supernatant and pelleted membrane fractions were tested for Cr(VI) reduction activity. The supernatant of the centrifugation showed almost the same Cr(VI) reduction activity as compared with that of the cell-free extract, indicating that the Cr(VI)-reducing activity of P. aeruginosa HP-14 is due to soluble enzyme. Moreover, the activity appeared to be the highest among the known activities, suggesting that the strain might be useful for remediation of Cr(VI)-contaminated sites.

• KSBB Journal
• /
• v.13 no.2
• /
• pp.162-167
• /
• 1998

The adsorption behaviors of two major cellulase components, endo I and exo II, from Trichoderma viride were investigated using $\alpha$-celluloses with different correlation crystallinity index(Cc) as substrates. The adsorption of cellulase enzyme components was significantly affected by the reaction condition and the physicochemical properties of the cellulose. The $\alpha$-cellulose was hydrolyzed in the presence of cellulase for various periods. The correlation crystallinity index of $\alpha$-cellulose increased with increasing the hydrolysis time. The adsorption was apparently found to obey the first-order kinetics, and the adsorption activation energy(Ea) was calculated from the adsorption rate constant(ka). The value of adsorption rate constant for endo I was larger than that of exo II. This means that endo I are adsorbed more rapidly than exo II. With the increase in correlation crystallinity index, the values of the adsorption rate constants for endo I and exo II decreased, respectively. The activation energy for the adsorption of exo II on the cellulose also was larger than that of endo I. Also adsorption activation energy of endo I and exo II increased with an increase in the crystallinity of sample cellulose.

• BMB Reports
• /
• v.38 no.3
• /
• pp.343-349
• /
• 2005

Gamma-glutamyltransferase (GGT, EC 2.3.2.2) which hydrolyzes glutathione (GSH), is required for the maintenance of normal intracellular GSH concentration. GGT is a membrane enzyme present in leukocytes and platelets. Its activity has also been observed in human neutrophils. In this study, GGT was purified from Triton X-100 solubilized neutrophils and its kinetic parameters were determined. For kinetic analyses of transpeptidation reaction, $\gamma$-glutamyl p-nitroanilide was used as the substrate and glycylglycine as the acceptor. Apparent $K_m$ values were determined as 1.8 mM for $\gamma$-glutamyl p-nitroanilide and 16.9 mM for glycylglycine. The optimum pH of GGT activity was 8.2 and the optimum temperature was $37^{\circ}C$. It had thermal stability with 58% relative activity at $56^{\circ}C$ for 30 min incubation. L-serine, in the presence of borate, was detected as the competetive inhibitor. Bromcresol green inhibited neutrophil GGT activity as a noncompetetive inhibitor. The neutrophils seem to contain only the isoenzyme that is present in platelets. We characterized the kinetic properties and compared the type of the isoenzyme of neutrophil GGT with platelet GGT via polyacrylamide gel electrophoresis (PAGE) under a standart set of conditions.

• Journal of Pharmaceutical Investigation
• /
• v.27 no.2
• /
• pp.99-108
• /
• 1997

To evaluate the feasibility of mucosal delivery of thyrotropin releasing hormone (TRH) through various mucosae, enzymatic degradation and stabilization of TRH in the nasal, rectal and duodenal extracts of rabbits were studied. TRH in the extracts was assayed by HPLC and its degradation was found to follow apparent first-order kinetics. The residual concentrations of TRH in the mucosal extracts of nasal, rectal and duodenal segments after 24 hr of incubation were found to be $65.1({\pm}1.1),\;19.7({\pm}2.7)$ and 0%, and in the serosal extracts, $65.6({\pm}5.5),\;75.2({\pm}1.1)$ and $68.7({\pm}1.4)%$, respectively. This result suggests that there is a significant difference in the activity of TRH-degrading enzymes among the sites of administration. The inhibition of TRH degradation in the mucosa extracts was kinetically investigated using various additives such as thimerosal, benzalkonium chloride, disodium edetate, ${\sigma}-phenanthroline$, dithiothreitol and dithioerythritol, and $IC_{50}$ values of inhibitors were calculated. The results obtained showed that thimerosal (0.5 mM) and benzalkonium chloride (0.141 mM) protected TRH from the enzymatic degradation in all the mucosa extracts more than 95% after 24 hr of incubation.

• Applied Chemistry for Engineering
• /
• v.19 no.3
• /
• pp.277-286
• /
• 2008

This paper deals with the theoretical derivation of the modified Haldane model of the substrate inhibition in the microbial growth processes. Based on the biological concepts of substrate-receptor complex working mechanisms, a new microbial kinetics of N-fold multiplex substrate inhibition and its generalization has been considered theoretically, which is natural expansion of the simple substrate inhibition mechanism in the enzyme reaction. As a result, the modified Haldane model of the substrate inhibition turns out to be a well-designed four-parameter kinetic model with a biological constant of the total substrate inhibition concentration.

• Bulletin of the Korean Chemical Society
• /
• v.10 no.6
• /
• pp.600-604
• /
• 1989

In order to find the rational methods for improving the thermal stability of subtilisin Carlsberg, the mechanisms of irreversible thermoinactivation of the enzyme were studied at $90^{\circ}C.$ At pH 4, the main process was hydrolysis of peptide bond. This process followed first order kinetics, yielding a rate constant of $1.26\;{\times}\;10^{-1}h^{-1}$. Hydrolysis of peptide bond of PMS-subtilisin occurred at various sites, which produced new distinct fragments of molecular weights of 27.2 KD, 25.9 KD, 25.0 KD, 22.3 KD, 19.0 KD, 17.6 KD, 16.5 KD, 15.7 KD, 15.0 KD, 13.7 KD, and 12.7 KD. Most of the new fragments originated from the acidic hydrolysis at the C-side of aspartic acid residues. However 25.0 KD, 15.7 KD, and 13.7 KD which could not be removed in purification steps stemmed from the autolytic cleavage of subtilisin. The minor process at pH 4 was deamidation at asparagine and/or glutamine residues and some extend of aggregation was also observed. However, the aggregation was main process at pH 7 with a first order kinetic constant of $16 h^{-1}.$ At pH 9, the main process seemed to be combination of deamidation and cleavage of peptide bond.