• 한국수산과학회지
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• 제41권4호
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• pp.243-247
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• 2008

The hemolysin of Vibrio cholerae non-O1 CT isolated from sea water was purified and characterized. The purified hemolysin displayed an optimum at $37^{\circ}C$ and exhibited more than 70% of residual hemolytic activity after incubation at $45^{\circ}C$ for 120 min. However, the activity dropped dramatically at temperature above $55^{\circ}C$. The purified protein showed the highest hemolytic activity at pH 7.0, while the activity was completely lost outside of the pH ranges of 5.0 and 10.0. The activity of hemolysin was inactivated by addition of divalent cations, such as $Cu^{2+},\;Fe^{2+},\;Hg^{2+},\;Mn^{2+},\;and\;Zn^{2+}$, however, the activity was not completely inhibited by additions of $Ca^{2+},\;Mg^{2+},\;K^+,\;Na^+,\;and\;Li^+$.

• BMB Reports
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• 제29권3호
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• pp.230-235
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• 1996

The interaction of ${\alpha}-ketoglutarate$ dehydrogenase complex (${\alpha}-KGDC$) with a hydrophobic fluorescent probe [1,1'-bi(4-aniline)naphthalene-5,5'-disulfonic acid] (bis-ANS) was studied. The punfied ${\alpha}-KGDC$ was potently inhibited by bis-ANS with an apparent half maximal inhibitory concentration ($IC_{50}$) of 9.8 ${\mu}m$ at pH 8.0. The catalytic activities of both the E1o and E2o subunits were predominantly inhibited while that of the E3 component was hardly affected. The binding of bis-ANS to the enzyme caused a marked enhancement and blue shift from 523 nm to 482 nm in the fluorescence emission spectrum. The dissociation constant ($K_d$) and the number of binding sites (n) were calculated to be 0.87 mM and 158, respectively. Allosteric regulators such as purine nucleotides and divalent cations further increased the fluorescence intensity of the $bis-ANS-{\alpha}-KGDC$ binary complex. These data suggest that the binding of these allosteric regulators to ${\alpha}-KGDC$ may cause the conformational changes in the enzyme and that bis-ANS could be used as a valuable probe to study the interaction of the multi-enzyme complex and its allosteric regulators.

• Parasites, Hosts and Diseases
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• 제56권1호
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• pp.81-86
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• 2018

Four isoforms of calcium binding proteins containing 2 EF hand motifs and a dynein light chain-like domain in the human liver fluke Opisthorchis viverrini, namely OvCaBP1, 2, 3, and 4, were characterized. They had molecular weights of 22.7, 21.6, 23.7, and 22.5 kDa, respectively and showed 37.2-42.1% sequence identity to CaBP22.8 of O. viverrini. All were detected in 2- and 4-week-old immature and mature parasites. Additionally, OvCaBP4 was found in newly excysted juveniles. Polyclonal antibodies against each isoform were generated to detect the native proteins in parasite extracts by Western blot analysis. All OvCaBPs were detected in soluble and insoluble crude worm extracts and in the excretory-secretory product, at approximate sizes of 21-23 kDa. The ion-binding properties of the proteins were analyzed by mobility shift assays with the divalent cations $Ca^{2+}$, $Mg^{2+}$, $Zn^{2+}$, and $Cu^{2+}$. All OvCaBPs showed mobility shifts with $Ca^{2+}$ and $Zn^{2+}$. OvCaBP1 showed also positive results with $Mg^{2+}$ and $Cu^{2+}$. As tegumental proteins, OvCaBP1, 2, and 3 are interesting drug targets for the treatment of opisthorchiasis.

• 미생물학회지
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• 제24권2호
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• pp.133-140
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• 1986

일산화탄소를 이용하여 자가영양적으로 성장한 Acinetobacter sp. 1 의 세포추출액은 혐 기성 실험조건하에서 thionin, methylene blue, 2,6-dichlorophenol-indophenol둥올 일산화탄소의 산회를 위한 전자수용체로 사용할 수 있었으나 NAD, NADP, FAD, 또는 FMN등은 천자수용체로 이용하지 못하였다. 이 세균에 존재하는 일산화탄소 산화효소는 유도효소로 밝혀졌고, pH 7.5와 $60^{\circ}C$에서 최대의 활성을 나타내었다. 이 효소의 활성화에너지는 6.1kcal/mol (25.5 kJ/mol)이며 일산화탄소에 대한 Km값은 $154{\mu}M$로 밝혀졌다. 그리고 잘 알려진 몇가지 금속 chelat tIng agent와 2가의 양이온들은 이 효소의 활성에 거의 영향을 미치지 않았는데 $Cu^{2+}$ 이온만은 이 효소의 활성을 완전히 억제시켰다. 또한 이 효소는 포도당과 숙신산에 의해 활성이 저해되었으며, hydrogenase의 활성도 나타내었다. 그리고 Acinetobacter sp. 1의 일산화탄소 산화효소는 Pseudomonas carboxydohydrogena의 일산화탄소 산화효소와 연역학적인 연관성이 없는 것으로 나타났다.

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

• Journal of Microbiology and Biotechnology
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• 제21권5호
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• pp.483-493
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• 2011

An acid phosphatase (HppA) activated by $NH_4Cl$ was purified 192- and 34-fold from the periplasmic and membrane fractions of Helicobacter pylori, respectively. SDS-polyacrylamide gel electrophoresis revealed that HppA from the latter appears to be several kilodaltons larger in molecular mass than from the former by about 24 kDa. Under acidic conditions (pH${\leq}$4.5), the enzyme activity was entirely dependent on the presence of certain mono- and/or divalent metal cations (e.g., $K^+$,$ NH_4{^+}$, and/or $Ni^{2+}$). In particular, $Ni^{2+}$ appeared to lower the enzyme's $K_m$ for the substrates, without changing $V_{max}$. The purified enzyme showed differential specificity against nucleotide substrates with pH; for example, the enzyme hydrolyzed adenosine nucleotides more rapidly at pH 5.5 than at pH 6.0, and vice versa for CTP or TTP. Analyses of the enzyme's N-terminal sequence and of an $HppA^-$ H. pylori mutant revealed that the purified enzyme is identical to rHppA, a cloned H. pylori class C acid phosphatase, and shown to be the sole bacterial 5'-nucleotidase uniquely activated by $NH_4Cl$. In contrast to wild type, $HppA^-$ H. pylori cells grew more slowly. Strikingly, they imported $Mg^{2+}$ at a markedly lowered rate, but assimilated urea rapidly, with a subsequent increase in extracellular pH. Moreover, mutant cells were much more sensitive to extracellular potassium ions, as well as to metronidazole, omeprazole, or thiophenol, with considerably lowered MIC values, than wild-type cells. From these data, we suggest that the role of the acid phosphatase HppA in H. pylori may extend beyond 5'-nucleotidase function to include cation-flux as well as pH regulation on the cell envelope.

• Journal of Microbiology
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• 제35권2호
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• pp.109-116
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• 1997

An extracellular proteinase of Candida albicans was purified by a combination of 0~75% ammonium sulfate precipitation, DEAE Sepharose Fast Flow ion exchange chromatography, and Sephacryl S-200 HR molecular sieve chromatography. Its mlecular weight was approximately 41 kDa on SDS-PAGE and isoelectric point was 4.4. The enzyme was inhibited by pepstain A. Optimum enzyme activity ranged from pH 2.0 to 3.5 with its maximum at pH 2.5 and a temperature of 45$^{\circ}C$. The addition of divalent cations, $Ca^{2+}$, Zn$^{2+}$ and $Mg^{2+}$, resulted in no significant inhibition of enzymatic activity. However, some inhibitory effects were observed by Fe$^{2+}$, Ag$^{2+}$ and Cu$^{2+}$. With BSA as substrate, an apparent $K_m$ was determined to be 7$\times$10$^{-7}$ M and $K_i$, using pepstatin A as an inhibitor, was 8.05$\times$10$^{-8}$ M. N-terminal amino acid sequence was QAVPVTLXNEQ. Degradation of BSA and fibronectin was shown but not collagen, hemoglobin, immunoglobulin G, or lysozyme. The enzyme preferred peptides with Glu and Leu at the P$_1$ position, but the enzyme activity was highly reduced when the P$_2$ position was phe or pro. This enzyme showed antigenicity against sera of patients with candidiasis.

• 청정기술
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• 제19권3호
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• pp.279-286
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• 2013

탄소원으로 구형 고분자수지 중 폴리스티렌-DVB 공중합으로 제조된 양이온교환수지를 사용하여 활성탄제조 시 활성탄의 주요물성인 비표면적과 비용적에 미치는 운전인자의 영향이 실험적으로 연구되었다. 활성탄의 비표면적과 비용적은 탄화공정에서 탄화온도 및 탄화시간이 증가함에 따라 감소하였고, 활성화공정에서는 활성화온도와 활성화시간에 따라 증가하였지만 특정 온도와 시간 이상에서는 감소하였다. 탄화공정에서 승온속도가 증가함에 따라 감소하였지만, 반대로 활성화 후 승온속도에 비례하는 물성을 보였다. 2가이온으로 포화된 수지로 제조된 활성탄은 새 수지로 제조된 활성탄에 비해 낮은 비표면적을 보였다.

• 한국식품영양과학회지
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• 제16권2호
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• pp.75-82
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• 1987

김치용 간절임 배추의 적당한 염도인 3%에 도달하는데 걸리는 간절임 시간은 5%에서는 l2시간 후에도 미달되었고, 10%는 약 7시간, 15%는 약3시간, 20%에서는 약 1시간 걸렸다. 절임중 잎은 줄기보다 빠르게 탈수되었는데 10시간 후에 잎과 줄기의 수분함랑은 각각 66와 64%로 낮아졌다. 생배추의 원부피는 간절임에 의해 59.90%가 감소하였고 생배추의 내부공간율은 35.06%나 감소하였다. 수침에 의한 제염속도는 염분침투속도보다 빨랐으며 약 7%수준에서 3%수준으로 되는데 잎은 약 1.2시간 들기는 약 3시간 걸렸다. 절임중 경도는 잎과 줄기에서 전체적으로 감소하였는데, 생체의 경우는 내부가 높았으나 간질임한 것은 점질부위의 경도가 증가하였다. 절임 과점에서 배추중의 $K^+,\;Ca^{2+},\;Mg^{2+}$ 금속이온은 감소한 반면 $Na^+$는 크게 증가했다. 절임과정과 수세과정에서 곰팡이, 효모, 세균의 수는 각각 58%, 40%, 45%줄었다.

• Parasites, Hosts and Diseases
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• 제46권1호
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• pp.37-40
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• 2008

Iron is an essential element for almost all living organisms. The possible role of iron for growth, adherence and cytotoxicity of Entamoeba histolytica was evaluated in this study. The absence of iron from TYI-S-33 medium stopped amebic growth in vitro. However, iron concentrations in the culture media of 21.4-285.6 ${\mu}M$ did not affect the growth of the amebae. Although growth was not retarded at these concentrations, the adhesive abilities of E. histolytica and their cytotoxicities to CHO cell monolayer were correlated with iron concentration. Amebic adhesion to CHO cell monolayers was significantly reduced by low-iron ($24.6{\pm}2.1%$) compared with $62.7{\pm}2.8\;and\;63.1{\pm}1.4%$ of amebae grown in a normal-iron and high-iron media, respectively. E. histolytica cultured in the normal- and high-iron media destroyed $69.1{\pm}4.3%\;and\;72.6{\pm}5.7%$ of cultured CHO cell monolayers, but amebae grown in the low-iron medium showed a significantly reduced level of cytotoxicity to CHO cells ($2.8{\pm}0.2%$). Addition of divalent cations other than iron to amebic trophozoites grown in the low-iron medium failed to restore levels of the cytotoxicity. However, when E. histolytica grown in low-iron medium were transferred to normal-iron medium, the amebae showed completely restored cytotoxicity within 7 days. The result suggests that iron is an important factor in the adherence and cytotoxicity of E. histolytica to CHO cell monolayer.