• Applied Chemistry for Engineering
• /
• v.31 no.5
• /
• pp.475-480
• /
• 2020

Chelates synthesized with Cu(II) ion and lactic acid or chitosan were applied to the hydrolysis of organophosphate simulant, DFP (diisopropyl fluorophosphate). Under the homogeneous reaction condition, Cu(II)-lactic acid chelate hydrolyzed DFP with the half life time of 37.1 min. Cu(II)-LMWS chitosan chelate was synthesized with 1 kDa molecular weight of chitosan, which showed low solubility, and then crystallized. The half life time for hydrolyzing DFP using Cu(II)-LMWS chitosan was 32.9 h indicating that the reaction rate is enhanced as much as 16 times more than that of using 18 kDa chitosan-Cu(II) complex. Under the homogeneous reaction condition, the half life time of Cu(II)-LMWS chitosan was 8.75 h. Therefore, we found out that the solubility of Cu(II)-LMWS chitosan makes the difference in the reaction rate as much as 4 times.

• Microbiology and Biotechnology Letters
• /
• v.18 no.4
• /
• pp.344-351
• /
• 1990

An alkalophilic microoganism producing a detergent-resistant alkaline protease was isolated from soil and identified as Baeiltus sp. The alkaline protease has been partially purified by ammonium sulfate fractionation, DEAE-Cellulose, CM-Cellulose and Sephdex G-100 column chromatography. The purified alkaline protease was highly active at pH 12-13 toward casein and stable at pH values from 6 to ll. The optimum temperature for the enzyme reaction was $55^{\circ}C$. The enzyme was completely inactivated by diisopropyl fluorophosphate (DFP) indicating that the enzyme was serine protease, but considerabiy stable in the presence of surface active agents.

• Applied Biological Chemistry
• /
• v.40 no.5
• /
• pp.388-394
• /
• 1997

Extracellular protease (bassiasin I), from the culture filtrate of entomopathogenic fungus Beauveria bassiana ATCC7159, was successively purified by precipitation with ammonium sulfate followed by DEAE-Sephadex A-50, CM-cellulose and Hydroxyapatite column chromatography. A typical procedure provided 41-fold purification with 13.6% yield. The molecular weight of the purified pretense (bassiasin I) was found to be approximately 32,000 by SDS-PAGE. Isoelectric-focusing analysis of the enzyme showed a pI of 9.5. $NH_2-terminal$ sequence of the pretense showed homology with those of the fungal proteases. The enzyme has an optimal pH for activity at 10.5 and is stable over pH 5.0-11.0. The maximum activity of the enzyme was at $60-65^{\circ}C$, and approximately 20% activity remained at $60^{\circ}C$ after 120 min. The pretense was inhibited by phenylmethylsulfonyl fluoride (PMSF) and diisopropyl fluorophosphate (DIPF).

• Proceedings of the Korean Society for Applied Microbiology Conference
• /
• 1986.12a
• /
• pp.532.2-532
• /
• 1986

Alkaline protease which is an important enzyme used in detergents, leather tanning and food industry was produced by alkalophilic bacterium, Bacillus sp. KCTC 1723 isolated from soil. The maximum productivity of the enzyme in alkaline medium containing 1% sodium bicarbonate was obtained by incubating for 3 days at 37$^{\circ}C$. The optimum pH of the enzyme was 11.5 and calcium ion was effective on stabilization of the enzyme at high temperature. The enzyme was not inhibited by metal chelating agent such as El)TA but inhibited by diisopropyl fluorophosphate. Purification of the enzyme was carried out DEAE- and CM-cellulose column chromatographies and molecular weight of the purified enzyme was determined

• Korean Journal of Microbiology
• /
• v.29 no.3
• /
• pp.167-171
• /
• 1991

An intracellular protease from cells of Pseudomonas carboxydohydrogena grown on nutrient broth was purified to better than 95% homogeneity in five steps using azocaseine as a substrate. The molecular weight of the native enzyme was determined to be 125, 000. Sodium dodecyl sulfate-gel electrophoresis revealedat least two non-identical subunits of molecular weight 70, 000 and 56, 000. The enzyme activity was completely ingibited by phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate. The enzyme was also inhibited by $Mg^{2+}$ , $Zn^{2+}$ , $Cd^{2+}$, $Cu^{2+}$ , and $Fe^{2+}$ , but was stimulated by iodoacetamide. Maximal reaction rate of the enzyme was observed at pH8.0 and 30.deg.C. The isoelectric point of the enzyme was found to be 7.5. The enzyme was unable to hydrolyze carbon monoxide dehydrogenase.

• BMB Reports
• /
• v.29 no.5
• /
• pp.455-461
• /
• 1996

A serine proteinase was purified from Acanthamoeba culbertsoni by 41~80% ammonium sulfate fractionation, ion exchange chromatography, affinity chromatography and gel filtration chromatography. The molecular weight of the purified enzyme was estimated to be 108.0 kDa by gel filtration chromatography and 54.0 kDa by SDS-PAGE. Therefore, the purified enzyme seemed to be a dimer. Isoelectric point was 4.5. The enzyme activity was highly inhibited by the serine proteinase inhibitors diisopropyl fluorophosphate (OFP) and phenylmethyl sulfonylfluoride (PMSF). It had a narrow pH optimum of 6.5~7.5 with a maximum at pH 7.0. These data suggested that the purified enzyme was a neutral serine proteinase. Optimal temperature was $37^{\circ}C$. It was stable for at least 16 h at $4^{\circ}C$ and $37^{\circ}C$, but it was rapidly inactivated at $65^{\circ}C$ The activity of the purified enzyme was not influenced significantly by $Mg^{2+}$, $Mn^{2+}$, $Zn^{2+}$ or $Ca^{2+}$. However, the enzyme activity was highly inhibited by $Hg^{2+}$ The enzyme degraded type I collagen and fibronectin, but not BSA, hemoglobin, lysozyme, immunoglobulin A or immunoglobulin G.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.24 no.5
• /
• pp.273-288
• /
• 1991

To elucidate the physiological and biochemical differences between chondrichthyes and osteichthyes, the properties of the specific digestive enzymes in cat-shark, Cephaloscyllium umbratile, and mackerel, Scomber japonicus, were studied. Homogenous trypsin proved through the disc-electrophoresis, SDS-PAG electrophoresis and gel filtration was obtained from the pancreas of cat-shark by $50-70\%$ saturated ammonium sulphate fractionation, DEAE-Sephadex A-50 column chromatography, benzamidine-Sepharose 6B affinity chromatography and Sephadex G-75-120 gel filtration. Two types of trypsins were also obtained from the pyloric caeca of mackerel by $30-70\%$ saturated ammonium sulphate fractionation and the slightly modified procedure from the method adopted in the purification of cat-shark trypsin. The two trypsins, designated trypsin A and B, were proved their homogeneity by disc- and SDS-PAG electrophoresis and gel filtration. The molecular weights of the trypsins were estimated to be 31,700 for cat-shark trypsin, 30,000 for mackerel trypsin A and 29,000 for mackerel trypsin B by SDS-PAG electrophoresis, but those were estimated to be 21,500 for cat-shark trypsin, 23,700 for mackerel trypsin A and 21,500 for mackerel trypsin B by gel filtration. The trypsins exhibited their optimum conditions at pH 9.0 and on temperature ranged from $45^{\circ}C\;to\;50^{\circ}C$ for cat-shark, and at pH 8.0 and a temperature of $50^{\circ}C$ for mackerel trypsin A and B, respectively. The cat-shark trypsin was stable at pH 10.0 and the temperature below $10^{\circ}C$, whereas the mackerel trypsin A and B, were stable in the range over pH 7.0 to pH 9.0 below $10^{\circ}C$ and at pH 8.0 below $35^{\circ}C$, respectively. The mackerel trypsins were severely inhibited by some heavy metal ions such as $Ag^{2+},\;Cu^{2+}\;and\;Hg^{2+}$ compared to cat-shark trypsin. All of the enzymes were also inhibited by antipain, leupeptin, TLCK(tosyllysine chloromethyl ketone) and SBTI(soybean trypsin inhibitor) remarkably. The inhibitory effects of PMSF(phenylmethane sulphonylfluoride), DFP(diisopropyl fluorophosphate) and benzamidine were indicated that these enzymes belong to serine-proteases.

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

• The Korean Journal of Nuclear Medicine
• /
• v.23 no.2
• /
• pp.219-223
• /
• 1989

Inflammation scan using radiolabelled leukocytes has high sensitivity and specificity. Several methods for labelling leukocytes have been evaluated using P-32 diisopropyl fluorophosphate (DFP-32), H-3 thymidine, Cr-51 chromate, Ga-67 citrate and Tc-99m-sulfur colloid. In-111-oxine has proved so far to be the most reliable agent for labelling leukocytes. In-111-oxine is, however, expensive, not easily available when needed, and its radiation dose to leukocytes is relatively high. Moreover, resolution of the resultant image is relatively poor. Tc-99m is still the agent of choice because of, as compared with the indium, its favorable physical characteristics, lower cost and availability. Now the technique for labelling the leukocytes with technetium is successfully obtained using the lipophilic HAPAO with higher efficiency for granulocytes than for other cells. With this technique it is possible to label leukocytes in plasma to improve the viability of the leukocytes. Inflammation scan using Tc-99m-HMPAO has been evaluated in several laboratories, and difference in methods for separation and labelling accounts for difference in efficiency, viability and biodistribution of the labelled leukocytes. We performed inflammation scan using leukocytes labelled with Tc-99m-HMPAO in three dogs 24 hours after inoculation of live E. Coli and A. Aureus in their right abdominal wall. We separated mixed leukocytes by simple sedimentation using 6% hetastarch (HES) and labelled the leukocytes with Tc-99m-HMPAO in 20% cell free plama diluted with phosphate buffer solution(Fig. 1). Uptake was high in the liver and spleen but is was minimal in the lungs on whole body scan. Kidneys and intestine showed minimal activity although it was high in the urinary bladder(Fig. 2). Uptake of labelled leukocytes in the inflammation site was do(mite on 2 hour-postinjection scan and abscess was clearly delineated on 24 hour-delayed scan with high target-to-nontarget ratio(Fig. 3, 4). Inflammation scan using mixed leukocytes labelled with Tc-99m-HMPAO is very sensitive and specific in early detection of inflammation.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.25 no.4
• /
• pp.282-290
• /
• 1992

Based on surplus production models using fishery data for the last 20 years, a stock assessment was conducted for the small yellow croaker in Korean waters. The maximum sustainable yields (MSY) from the Schaefer and Fox models were estimated to be 37,000 metric tons (mt) and 33,450 mt. Zhang's model using time-series biomass with instantaneous coefficients of fishing mortality (F) and using time-series biomass and catch yielded MSY estimates of 45,328 mt and 40,160 mt, respectively. A yield-per-recruit analysis showed that the current yield per recruit of about 20g with F= 1.11 $yr^{-l}$, where the age at first capture $(t_c)$ is 0.604, was much lower than the maximum possible yield per recruit of 43g. Fixing $t_c$ at the current level and reducing fishing intensity (F) from 1.11 $yr^{-l}$ to 0.4 $yr^{-l}$ yielded only a small increase in predicted yield per recruit, from 20 to 25g. However, estimated yield per recruit increased to 43g by increasing $(t_c)$ from the current age (0.604) to age three with F fixed at the current level. This age at first capture corresponded to the optimal length which was obtained from the $F_{0.1}$ method. According to the analysis of stock recovery strategies employing the Zhang model, the optimum equilibrium biomass $(B^*_{MSY})$ which produces the maximum yield could be achieved after approximately five years at the lower fishing intensity (F=0.5).