• Parasites, Hosts and Diseases
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• v.33 no.4
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• pp.323-330
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• 1995

Protease activity was identified in crude extracts of Pnrqgonimw westermnni eggs which were purified from infected dog lungs, isolated on 14 weeks after metacercarial challenge. The eggs were used after removing possibly contaminated host or worm tissues on their shell surfaces. In the crude egg extracts, high proteolytic activities against carboBfrb enzoyl - ph enylalanyl - arginyl-4- methoxy- β- naphthylamide (Cbz - phe - arg- MNA) and Azocoll were detected whereas those against succinyl-alanyl-propyl-phenylalanyl-p- nitroanilide (Suc-ala-pro-phe-pNA) were not revealed. The eVe eBdlibited the maximal activity at pH 6. Its activity was inhibited by specific cysteine protease inhibitors, 105 M I- trans-epoxysuccinylleucylamido (4-guanidino) butane (E-64) and 1 mM iodoacetamide (LAA) while potentiated by 6.5-fold in the presence of 2.5 mM dithiothreitol (DTT) . When the enzyme was purified partially by Sephacryl S-300 High Resolution gel filtration, it migrated as a single homogeneous band at 35 kDa. The 35 kDa cysteine protease has been recognized neither in the metacercariae nor in the adult. These findings indicated the presence of at least one protease of cathepsin family in immature eggs of f westernani.

• Journal of the Korean Chemical Society
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• v.50 no.5
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• pp.362-368
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• 2006

SH group modifying chemicals were used to characterize the eight cysteine residues of cabbage PLD. 5,5-dithiobis(2-nitrobenzoate)(DTNB) was used to titrate the SH group of cysteine residues . Based on the optical density at 412nm due to the reduced DTNB, 4 SH groups are found to be present in a native PLD while 8 SH groups in the denatured PLD whose tertiary structure was perturbed by 8M urea. The results imply that among the 8 cysteine residues of PLD, the half(4) are exposed on the surface whereas the other half are present at the interior of the enzyme tertiary structure. The PLD was inactivated by SH modifying reagents such as p-chloromercuribenzoate(PCMB), iodoacetate, iodoacetamide, and N-ethylmaleimide. At the addition of dithiothreitol(DTT) only the PCMB inhibited PLD activity was recovered reversibly. The micro-environment of the exposed SH group of cysteine residues was examined with various disulfide compounds with different functional groups and we found that anionic or neutral disulfides appear to be more effective than the positively charged cystamine for inactivating the PLD activity. The effect of redox state of cysteine residues on the PLD activity was further explored with H2O2. The oxidation of SH groups by H2O2 inhibited the PLD activity more than 70%, which was mostly recovered by DTT. From these results, we could confirm chemically that all the cysteine residues of PLD are present as in their reduced SH forms and the 4 SH groups exposed on the surface of the enzyme may play important roles in the regulation of PLD activity.

• Korean Journal of Food Science and Technology
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• v.36 no.3
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• pp.440-447
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• 2004

Characteristics of human ${\beta}-carotene$ 15,15'-dioxygenase isolated by recombinant DNA technology was elucidated. Optimal pH and temperature were 9.0 and $40^{\circ}C$, respectively. Enzyme activity was temperature-sensitive. Enzyme was stable at pH 6.0-9.0 for 24 hr and under $5^{\circ}C$. Half-life of enzyme at $35^{\circ}C$ was 40 min. Crude preparations of enzyme were inhibited by ferrous ion-chelating agent and sulfhydryl-binding agent. GSH offsets inhibitory effect of PCMB. With increasing substrate concentrations, enzyme activity gave typical Michaelis-Menten curve, Based on Hanes-Woolf plot of data, $K_{m}\;and\;V_{max$ were $3.39{\times}10^{6}\;M\;and\;1.2\;pmol/mg$ protein/min, respectively.

• Journal of Microbiology
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• v.43 no.3
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• pp.285-294
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• 2005

A bacterial strain M4-7 capable of degrading various polyesters, such as poly$(\varepsilon-caprolactone)$, poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3-hydroxyoctanoate), and poly(3-hydroxy-5-phenylvalerate), was isolated from a marine environment and identified as Pseudomonas alcaligenes. The relative molecular mass of a purified extracellular medium-chain-length poly(3-hydroxyalkanoate) (MCL-PHA) depolymerase $(PhaZ_{palM4-7})$ from P. alcaligenes M4-7 was 28.0 kDa, as determined by SDS-PAGE. The $PhaZ_{palM4-7}$ was most active in 50 mM glycine-NaOH buffer (pH 9.0) at $35^{\circ}C$. It was insensitive to dithiothreitol, sodium azide, and iodoacetamide, but susceptible to p-hydroxymercuribenzoic acid, N-bromosuccinimide, acetic anhydride, EDTA, diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride, Tween 80, and Triton X-100. In this study, the genes encoding MCL-PHA depolymerase were cloned, sequenced, and characterized from a soil bacterium, P. alcaligenes LB19 (Kim et al., 2002, Biomacro-molecules 3, 291-296) as well as P. alcaligenes M4-7. The structural gene $(phaZ_{palLB19})$ of MCL-PHA depolymerase of P. alcaligenes LB19 consisted of an 837 bp open reading frame (ORF) encoding a protein of 278 amino acids with a deduced $M_r$ of 30,188 Da. However, the MCL-PHA depolymerase gene $(phaZ_{palM4-7})$ of P. alcaligenes M4-7 was composed of an 834 bp ORF encoding a protein of 277 amino acids with a deduced Mr of 30,323 Da. Amino acid sequence analyses showed that, in the two different polypeptides, a substrate-binding domain and a catalytic domain are located in the N-terminus and in the C-terminus, respectively. The $PhaZ_{palLB19}$ and the $PhaZ_{palM4-7}$ commonly share the lipase box, GISSG, in their catalytic domains, and utilize $^{111}Asn$ and $^{110}Ser$ residues, respectively, as oxyanions that play an important role in transition-state stabilization of hydrolytic reactions.

• International Journal of Oral Biology
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• v.42 no.4
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• pp.155-161
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• 2017

Teeth and bones are highly mineralized tissues containing inorganic minerals such as calcium phosphate, and a growing number of evidences show that their mineral content is associated with many diseases. Although the quantification of mineral contents by micro-computed tomography(micro- CT) has been used in diagnosis and evaluation for treating bone diseases, its application for teeth diseases has not been well established. In this study, we attempted to estimate a usefulness of a high-resolution micro-CT in analysis of human teeth. The teeth were scanned by using the Skyscan 1172 micro-CT. In order to measure tooth mineral content, beam hardening effect of the machine was corrected with a radiopaque iodine-containing substance, iodoacetamide. Under the maximum resolution of $6.6{\mu}m$, X-ray densities in teeth and hydroxyapatite standards were obtained with Hounsfield unit (HU), and they were then converted to an absolute mineral concentration by a CT Analyzer software. In enamel layer of cusp area, the mean mineral concentration was about $2.14mg/mm^3$ and there was a constant mineral concentration gradient from the enamel surface to the dentinoenamel junction. In the dentin of middle 1/3 of tooth, the mean mineral concentration was approximately $1.27mg/mm^3$ and there was a constant mineral concentration gradient from the outer of root to the pulp side, ranging from 1.3 to $1.06mg/mm^3$. In decay region of dentin, the mineral content was gradually decreased from the intact inner side to the decayed surface. These results suggest that high-resolution micro-CT can be as a useful tool for non-invasive measurement of mineral concentration in teeth.

• Korean Journal of Environmental Agriculture
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• v.14 no.3
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• pp.329-337
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• 1995

The effect of N-(4-chlorophenyl) maleimide(CPMI), plant growth regulators, and alkylating agents on gluathione(GSH) content and glutathione S-transferase(GST) activity was examined with 3-day-old etiolated sorghum(Sorghum bicolor [L.] Moench) seedlings. The GSH content and GST activity of untreated seedlings were higher in shoots than that in roots. Response of GST activity in coleoptile was significantly greater than in other tissues of sorghum seedling. In CPMI-treated seedlings, GSH content was not significantly different from that in untreated seedlings. CPM treatment resulted in 2.3-fold increase in GST activity measured with metolachlor as substrate in the coleoptile region. In contrast, change in GST activity measured with metolachlor as substrate in the coleoptile region. In contrast, change in GST activity measured with 1-chloro-2, 4-dinitrobenzene did not occur. The increase of GST activity was caused by induction of a GST isozyme, which is substrate-specific to metolachlor. Subsequently, two hypotheses related to metolachlor detoxification were evaluated on the basis of regulation of plant growth regulators and substrate induction of GST activity. In coleoptile, GST activity measured with metolachior was increased to 2.1-and 3.4-fold by both 2, 4-dichlorophenoxyacetic acid(2,4-D) and metolachlor treated at the germination stage of sorghum, respectively. Treatments of 2.4-D and metolachlor also induced isozymes exhibiting the activity toward metolachlor. One of the isozymes was co-eluted with that induced by CPMI. These results indicated that increase in GST activity by CPMI may be partially related to auxin regulation and substrate induction.