• Journal of Microbiology and Biotechnology
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• v.9 no.3
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• pp.249-254
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• 1999

A catechol 2,3-dioxygenase (C23O) was purified to apparent homogeneity from Pseudomonas putida SU10 through several purification steps consisting of ammonium sulfate precipitation and chromatographies on DEAE 5PW, Superdex S-200, and Resource-Q. Gel filtration indicated a molecular mass under nondenaturing conditions of about 130 kDa. The enzyme has a subunit of 34 kDa as was determined by SDS-PAGE. These results suggest that the native enzyme is composed of four identical subunits. The N-terminal amino acid sequence (30 residues) of the enzyme has been determined and exhibits high identity with other extradiol dioxygenases. The reactivity of this enzyme towards catechol and methyl-substituted catechols is somewhat different from that seen for other catechol 2,3-dioxygenases, with 4-methylcatechol cleaved at a higher rate than catechol or 3-methylcatechol. $K_m$ values of the enzyme for these substrates are between 3.5 and 5.7 M.

• Journal of the Korean Society of Food Science and Nutrition
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• v.29 no.4
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• pp.719-725
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• 2000

This study focused on the purification and characterization of ACE inhibitor from Porphyra yezoensis. The dried Porphyra yezoensis was ground and hydrolyzed with 2.5 N HCl, followed by neutralization and centrifugation. Then, the subsequential purification of ACE inhibitor was carried out by Amberlite XAD 8, DEAE-Toyopearl 650C, Sephadex LH-20 column chromatography and reverse phase HPLC with C18 column. The purified ACE inhibitor was peptide which consisted of glycine (24.5%), arginine (56.8%) and proline (18.8%). Also, it showed the competitive inhibition pattern to ACE. The apparent molecular mass of purified peptide was 580 dalton, and an IC50 value of ACE inhibitor was 10.6 $\mu\textrm{g}$.

• Proceedings of the Korean Nutrition Society Conference
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• 2001.05a
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• pp.313.1-313
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• 2001

• BMB Reports
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• v.40 no.3
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• pp.412-418
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• 2007

Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is an unique antioxidant enzyme that directly reduces lipid hydroperoxides in biomembranes. In the present work, the entire encoding region for Oryza sativa PHGPx was expressed in Escherichia coli M15, and the purified fusion protein showed a single band with 21.0 kD and pI = 8.5 on SDS- and IFE-PAGE, respectively. Judging from CD and fluorescence spectroscopy, this protein is considered to have a well-ordered structure with 12.2% $\alpha$-helix, 30.7%$\beta$-sheet, 18.5% $\delta$-turn, and 38.5% random coil. The optimum pH and temperature of the enzyme activity were pH 9.3 and 27$^{\circ}C$. The enzyme exhibited the highest affinity and catalytical efficiency to phospholipid hydroperoxide employing GSH or Trx as electron donor. Moreover, the protein displayed higher GSH-dependent activity towards t-Butyl-OOH and $H_2O_2$. These results show that OsPHGPx is an enzyme with broad specificity for hydroperoxide substrates and yielded significant insight into the physicochemical properties and the dynamics of OsPHGPx.

• Microbiology and Biotechnology Letters
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• v.21 no.6
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• pp.588-593
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• 1993

Xylanase(1, 4-beta-D-xylan xylanohydrolase` EC 3.2.1.8) II was purified from Penicillium verruculosum by using the techniques of two anion exchange chromatographies, and gel filtration. The molecular weight of this enzyme was about 22, 000 as determined by SDS-electrophoresis. The enzyme showed hydropytic activity toward xylan but did not catalyze hydrolysis of Rho-nitrophenyl-beta-D-xylopyranoside, Rho-nitrophenyl-beta-D-glucopyranoside, Rho-nitrophenyl-beta-D-cellobiopyranoside, and celluloses such as Avicel, cotton, filter paper, carboxymethylcellulose.

• Journal of Microbiology
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• v.35 no.2
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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.

• Proceedings of the Korean Society of Life Science Conference
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• 2005.04a
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• pp.116-116
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• 2005

• Journal of Microbiology and Biotechnology
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• v.14 no.4
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• pp.863-867
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• 2004

$\alpha$-Galactosidase from A. oryzae DR-5 was induced in the presence of melibiose, raffinose, galactose, and locust bean galactomannan. The enzyme was purified to homogeneity by precipitation with acetone followed by ion-exchange chromatography using DEAE-Sephacel. The purified enzyme showed a single band in both nondenaturing-PAGE and SDS-PAGE. The enzyme was a glycoprotein in nature by activity staining. The molecular weight of the purified enzyme was 93-95 kDa by SDS-PAGE. The enzyme exhibited the optimum pH and temperature at 4.7 and $60^\circ{C}$, respectively. $\alpha$-Galactosidase activity was strongly inhibited by $Ag^{2+}, Hg^{2+}, Cu^{2+}$, and galactose. EDTA, 1,10-phenanthraline, and PMSF did not inhibit the enzyme activity, whereas N-bromosuccinimide completely inhibited enzyme activity. Investigation by TLC showed complete hydrolysis of stachyose and raffinose in soymilk in 3 h at pH 5.0 and $50^\circ{C}$.

• Journal of Microbiology and Biotechnology
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• v.20 no.12
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• pp.1681-1688
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• 2010

A screening for cellobiohydrolase (CBH) activity was performed and Fomitopsis pinicola KMJ812 was selected for further characterization as it produced a high level of CBH activity. An extracellular CBH was purified to homogeneity by sequential chromatography of F. pinicola culture supernatants. The molecular mass of the F. pinicola CBH was determined to be 64 kDa by SDS-PAGE and by size-exclusion chromatography, indicating that the enzyme is a monomer. The F. pinicola CBH showed a $t_{1/2}$ value of 42 h at $70^{\circ}C$ and catalytic efficiency of $15.8mM^{-1}s^{-1}(k_{cat}/K_m)$ for p-nitrophenyl-${\beta}$-D-cellobioside, one of the highest levels seen for CBH-producing microorganisms. Its internal amino acid sequences showed a significant homology with hydrolases from glycoside hydrolase family 7. Although CBHs have been purified and characterized from other sources, the F. pinicola CBH is distinguished from other CBHs by its high catalytic efficiency and thermostability.

• Journal of Microbiology and Biotechnology
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• v.15 no.4
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• pp.689-693
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• 2005

Bacillus sp. JB-99, when grown in a chemically defined medium containing lactose as a carbon source, yielded 3,860 U/ml extracellular $\beta$-mannanase, which was high compared to other examined carbon sources. Among the nitrogen sources, yeast extract enhanced the enzyme activity. The enzyme production was growth-associated. The enzyme was optimally active at $65^{\circ}C$, pH 10, and had a half-life of 190 min at $65^{\circ}C$. N-Bromosuccinamide and $AgNO_3,\;CuSO_4$, and $HgCl_2$ strongly inhibited the enzyme, whereas $Ca^{2+}$ stimulated the enzyme activity. The $\alpha$-galactosidase enzyme production was not found in any of the enzyme assays.