• Microbiology and Biotechnology Letters
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• v.16 no.4
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• pp.310-315
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• 1988

Two isoenzymes of chorismate mutase(E.C.5.4.99.5) designated as chorismate mutase I(CM I) and chorismate mutase II(CM II), were detected and partially purified from a sp. of intrasporangium isolated from soil. CM I and CM II had pH optima of pH 6.5 and 8.0, respectively and showed the same temperature optimum of 45$^{\circ}C$. The activation energy of the enzymatic reaction was estimated to be 14.7kcal/ mole with CM I and 10.8kcal/mole with CM II. The affinity of isoenzyme CM I for substrate(Km= 1.35mM) was almost the same level as that of CM II(Km = 1.22mM). Both isoenzymes were stable at pH values ranged from pH 6.5 to 9.0, but rapidly denaturated at temperatures above 45$^{\circ}C$. CM II was activated about 7$^{\circ}C$ of its activity by $Ba^{++}$ or $Mg^{++}$ while CM I was slightly inhibited by the same metal ions. Thiol compounds were found not to be necessary for stability of the two enzymes but Co$^{++}$ and EDTA had a little stabilizing effect on CM II only. p-Chloromercuribenzoate strongly inactivated the activities of both enzymes but the reducing agents such as dithiothreitol and L-cysteine protected them against the pCMB inhibition.

• Korean Journal of Microbiology
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• v.30 no.4
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• pp.310-315
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• 1992

Absorption of $\textrm{Pb}^{2-}$ from aqueous solution was studied using Staphylococcus epidermidis. Cells of exponential phase were employed as absorbents. Uptake ratio defined as the ratio of amount of $\textrm{Pb}^{2+}$ absorbed to that of initial $\textrm{Pb}^{2+}$. Absorption of $\textrm{Pb}^{2+}$ increased with increase in cell concentration. while amount of $\textrm{Pb}^{2+}$ per unit cell mass decreased. Uptake ratio of $\textrm{Pb}^{2+}$ augmented and then diminished after exhibiting a maximum as the pH of the solution increased. Equilibrium absorption of $\textrm{Pb}^{2+}$ deviated from Freundlich isotherm especially at higher concentration of $\textrm{Pb}^{2+}$ due to the precipitation phenomena. HCI and EDTA were founded to desorb $\textrm{Pb}^{2+}$ more effectively than $\textrm{Na}_{2}\textrm{CO}_{3}$ or $\textrm{NaHCO}_{3}$. After 10 cycles of absorption and desorption. $\textrm{Pb}^{2+}$ absorption capability remained almost unchanged and the biomass had leaked out 30-40 wt/%. Uptake ratio of Pb2+ decreased in the presence of other heavy metal ions due to the competitive absorption The inhibition of $\textrm{Pb}^{2+}$ absorption appeared to have a strong correlation with ionic radius of the competing ions. Especially $\textrm{Cr}^{3-}$, $\textrm{Co}^{2+}$ or $\textrm{Fe}^{2+}$ having smaller ionic radius depressed more significantly the uptake of $\textrm{Pb}^{2+}$ than any other metal ions tested.

• Journal of the Korean Society of Food Science and Nutrition
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• v.15 no.2
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• pp.158-164
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• 1986

Polyphenol oxidase in apple (Golden Delicious) was extracted, partially purified and its properties were found as follows; Polyphenol oxidase showed optimum pH for activity at 6.5 and optimum temperature at $30^{\circ}C$ and high affinity to o-diphenol compounds. Cysteine, ascorbic acid and sodium metabisulfite appeared to be most effective inhibitors. EDTA showed a slight inhibition. During the enzyme was kept in test tube at $4^{\circ}C\;and\;20^{\circ}C$ for a week, polyphenol oxidase activity decreased sharply during the first four days at $20^{\circ}C$, then decreased slowly as the storage was prolonged. At $4^{\circ}C$, the polyphenol oxidase activity appeared to be relatively stable during the first two days before activity began to decline sharply. Polyacrylamide disc gel electrophoresis indicated four bands with polyphenol oxidase activity. Three bands and one band of the active bands were observed after heating for 1hr at $60^{\circ}C\;and\;70^{\circ}C$ respectively. The enzyme activity was observed 40% after treatment at $60^{\circ}C$ and 5% after treatment at $70^{\circ}C$. Therefore, no difference in the thermal stability was observed between active bands and the enzyme activity.

• Korean Journal of Clinical Laboratory Science
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• v.53 no.1
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• pp.49-59
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• 2021

This study investigated the use of real-time PCR melting curves for the diagnosis of blaKPC and blaNDM genes among the most frequently detected carbapenemase-producing Enterobacteriaceae in Korea. As a means of addressing the shortcomings of phenotype tests and conventional PCR. The modified Hodge test confirmed positivity in 25 of 35 strains, and carbapenemase inhibition testing confirmed positivity in 14 strains by meropenem+PBA or meropenem+EDTA. PCR analysis showed amplification products in 25 strains of Klebsiella pneumoniae carbapenemases (KPC), 10 of K. pneumoniae, 5 of E. coli, 5 of A. baumannii, 4 of P. aeruginosa, and 1 of P. putida. New Delhi metallo β-lactamase (NDM) identified amplification products in 8 strains, that is, 2 K. pneumoniae, 3 E. coli, 1 P. aeruginosa, 1 E. cloacae, and 1 P. retgeri strains. Real-time PCR melting curve analysis confirmed amplification in 25 strains of KPC and 8 strains of NDM, and these results were 100% consistent with PCR results. In conclusion, our findings suggest early diagnosis of carbapenem resistant Enterobacteriaceae by real-time PCR offers a potential means of antibacterial management that can prevent and control nosocomial infection spread.

• Journal of Microbiology and Biotechnology
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• v.24 no.2
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• pp.245-253
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• 2014

A fibrinolytic enzyme was produced by an edible mushroom of Pleurotus ostreatus using submerged culture fermentation. The enzyme was purified from the culture supernatant by applying a combination of freeze-thaw treatment, ammonium sulfate precipitation, hydrophobic interaction, and gel filtration chromatographies. The enzyme was purified by a 147-fold, with a yield of 7.54%. The molecular masses of the enzyme an determined by gel filtration and SDS-PAGE were 13.6 and 18.2 kDa, respectively. The isoelectric point of the enzyme was 8.52. It hydrolyzed fibrinogen by cleaving the ${\alpha}$ and ${\beta}$ chains of fibrinogen followed by the ${\gamma}$ chains, and also activated plasminogen into plasmin. The enzyme was optimally active at $45^{\circ}C$ and pH 7.4. The enzyme activity was completely inhibited by EDTA, whereas protease inhibitors of TPCK, SBTI, PMSF, aprotinin and pepstatin showed no inhibition on its activity. The partial amino acid sequences of the enzyme as determined by Q-TOF2 were ATFVGCSATR, GGTLIHESSHFTR, and YTTWFGTFVTSR. These sequences showed a high degree of homology with those of metallo-endopeptidases from P. ostreatus and Armillaria mellea. The purified enzyme can also be applied as a natural agent for oral fibrinolytic therapy or prevention of thrombosis.

• The Korean Journal of Mycology
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• v.28 no.1
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• pp.60-65
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• 2000

Two fibrinolytic enzymes were purified from the fruiting bodies of Tricholoma saponaceum. The enzymes have a molecular weight of 18(FE-1) and 18.2(FE-2) kDa, respectively, and include $Zn^{2+}$ ion as determined by ICP/MS. The N-terminal amino acid sequence of the two enzymes were exactly the same: A-L-Y-V-G-X-S-P-X-Q-Q-S-L-L-V. The activity of FE-1 was highly inhibited by EDTA and 1,10-phenanthroline, indicating that the enzyme is a metalloprotease. The activity of FE-1 was slightly increased by $Mg^{2+},\;Zn^{2+},\;Fe^{2+}\;and\; Co^{2+}$, however, the enzyme activity was totally inhibited by $Hg^{2+}$. Addition of $Zn^{2+}\;and\;Co^{2+}$ reversed the inhibition caused by 1,10-phenanthroline. It has a pH optimum at pH 7.5, suggested that FE-1 was a neutral protease. It shows the maximum fibrinolytic activity at $55^{\circ}C$, is completely inactivated above at $65^{\circ}C$.

• The Korean Journal of Mycology
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• v.36 no.2
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• pp.183-188
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• 2008

A fibrinolytic serine protease was purified from the fruiting bodies of an edible mushroom, Albatrellus confluens. The enzyme had a molecular mass of 30086.41 Da, as measured by MALDI-TOF mass spectrometry. The N-terminal amino acid sequence of the enzyme was Glu-Thr-Val-Thr-Glu-Thr-Thr-Ala -Pro-Trp-Gly-Leu-Ser-Arg-Ile. It displayed optimal activity at $50^{\circ}C$ and within a pH range of $8.0{\sim}10.0$, suggesting that the enzyme is an alkaline protease. The enzyme was stable up to $30^{\circ}C$. The enzyme displayed a strong substrate specificity for the synthetic peptide, N-Suc-Ala-Ala-Pro-Phe pNA. The enzyme activity was completely inhibited by addition of PMSF, indicating that the enzyme is a serine protease. No inhibition was observed following addition of E-64, pepstatin, or EDTA. The activity of the purified enzyme was decreased in the presence $Fe^{2+}$ or $Co^{2+}$, and the enzyme was completely inhibited by addition of $Hg^{2+}$. From these results, we propose that Albatrellus confluens could be used for biofunctional foods development and has potential therapeutic value for the treatment of vascular diseases.

• Microbiology and Biotechnology Letters
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• v.41 no.1
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• pp.60-69
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• 2013

Purified melanin pigment from Klebsiella sp. GSK was characterized by thermogravimetric, differential thermal, X-ray diffraction and elemental analysis. This melanin pigment is structurally amorphous in nature. It is thermally stable up to $300^{\circ}C$ and emits a strong exothermic peak at $700^{\circ}C$. Its carbon, hydrogen and nitrogen composition is 47.9%, 6.9% and 12.0%, respectively. It was used to scavenge metal ions and free radicals. After immobilizing the pigment and using it to adsorb copper and lead ions, the metal ion adsorption capacity was evaluated by atomic absorption spectroscopy (AAS) and the identity of melanin functional groups involved in the binding of metal ions was determined by Fourier transform infrared (FT-IR) spectroscopy. Batch adsorption studies showed that 169 mg/g of copper and 280 mg/g of lead were adsorbed onto melanin-alginate beads. The metal ion adsorption capacity of the melanin-alginate beads was relatively significant compared to alginate beads. The metal ion desorption capacity of HCl was greater (81.5% and 99% for copper and lead, respectively) than that of EDTA (80% and 71% for copper and lead, respectively). The ability of the melanin pigment to scavenge free radicals was evaluated by inhibition of the oxidation of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and was shown to be about 74% and 98%, respectively, compared with standard antioxidants.

• Journal of the Korean Chemical Society
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• v.24 no.1
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• pp.25-33
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• 1980

Pyrocatechase as a phenolytic dioxygenase was extracted from the benzoate-induced cells of a soil bacterium, a member of Pseudomonadaceae, and purified partially by DEAE-cellulose ion-exchange chromatography and Sephadex G-75 gel filtration. Final preparation of the enzyme yielding 200 fold purification over the crude extracts showed a specific activity of about 40 ${\mu}moles$ per minute per mg protein based on catechol as the substrate. The enzyme showed a very limited substrate specificity towards catechol for its catalytic activity. Based on the inhibition study with the substrate analogues, it was assumed that ortho dihydroxy groups on the aromatic ring may participate in the enzyme-substrate binding. The $K_m$ value for catechol was obtained as $1.9{\times}10^{-6}M$, and the optimum activity of the enzyme was obtained at the pH range of 7∼10 and $35^{\circ}C$. With SH-group blocking agents the enzyme was inhibited seriously. The activity of enzyme was also inhibited by the addition of some heavy metals, $Ag^+$ and $Cu^{2+}$, but was not affected by EDTA. General property of the enzyme was characterized and the possible nature of the enzyme active center was also discussed.

• Korean Journal of Veterinary Research
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• v.44 no.3
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• pp.455-462
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• 2004

Change in ${\beta}$-tubulin nucleic acid and protein sequences was the only known difference between Haemonchus contortus fenbendazole (FBZ)-resistant and -susceptible isolates. This change was sufficient to determine the pathologic effect induced by FBZ treatment. This research was initiated to investigate further differences from these two isolates. Since ${\beta}$-tubulin is involved in formation of microtubule, which has functions in secretory vesicle transport, DNase activities from excretory/secretory products (ESP) of the two isolates were compared, based on pH, sensitivity to DNase inhibitors, molecular masses and production of 3'-OH. The most significant difference detected was that a 38.5 kDa DNase activity was identified from ESP of H. contortus FBZ-susceptible isolates but not from those of H. contortus FBZ-resistant isolates. However, it was shown that the 38.5 kDa DNase is expressed with similar level of activity in intestine and whole worm of H. contortus FBZ-resistant and -susceptible isolates. This result demonstrated that the secretory transport pathway of the 38.5 kDa DNase was inhibited by unknown mechanisms, which may be related with ${\beta}$-tubulin sequence change in FBZ-resistant isolates. Other DNases of 34, 36 and 37 kDa were detected from ESP of both H. contortus FBZ-resistant and -susceptible isolates. Overall DNase activities found from ESP of these two isolates were not inhibited by 10 mM EDTA at pH 5.0, but largely inhibited by pH 7.0. In addition, DNase activities in two isolates produced DNA fragments with mixtures of 3'- hydroxyls (OH) and 3'-phosphates (P) at each pH although the 3'-end labeling ratios at pH 5.0 and 7.0 were shown different. Identification of inhibition of the 38.5 kDa DNase secretion in FBZ-resistant isolates suggests existence of further differences, in addition to ${\beta}$-tubulin sequence change, in two isolates. This shows complex effect of FBZ on H. contortus biological mechanisms.