• KSBB Journal
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• v.23 no.6
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• pp.489-493
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• 2008

Although the seafood cooking drips were the byproducts from the fishery industry and being wasted, it had many nutrients including proteins. In this study, the effect of a gamma irradiation on the cooking drips from Hizikia fusiformis, Enteroctopus dofleni and Thunnus thynnus were investigated. The cooking drips were extracted with 70% ethanol solution, and the extracts were analysed for the protein concentration by three different methods of Lowry, BCA and Kjeldahl. The extracts were irradiated with different doses and the protein contents were compared with respect to the absorbed doses. Total content of the proteins was increased with increasing irradiation dose. The change of protein pattern in the irradiated cooking drips was also confirmed by SDS-PAGE analysis. These results shown that the proteins in cooking drips could be unfolded or aggregated by the irradiation. Therefore, gamma irradiation could be considered as an effective method for extracting useful proteins.

• Korean Journal of Environmental Biology
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• v.39 no.4
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• pp.445-451
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• 2021

In this study, we described the production of an antibody to living modified organisms (LMOs) containing the gene encoding for 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) from Agrobacterium tumefaciens strain CP4 EPSPS provides resistance to the herbicide glyphosate (N- (phosphonomethyl) glycine). These LMOs were approved and have recently been used in the feed, food production, and processing industries in South Korea. Highly efficient monoclonal antibody (mAb) production is crucial for developing assays that enable the proper detection and quantification of the CP4 EPSPS protein in LMOs. This study describes the purification and characterization of recombinant CP4 EPSPS protein in E. coli BL21 (DE3) based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrixassisted laser desorption/ionization time-of-flight mass spectrometry. The production of mAbs was undertaken based on the standard operating procedure of Abclon, Inc.(South Korea), and the purity of the mAbs was assessed using SDS-PAGE. The following five mAb clones were produced: 2F2, 4B9, 6C11, 10A9, and 10G9. To verify the efficiency and specificity of the five developed mAbs, we performed Western blotting analysis using the LM (living modified) cotton crude extracts. All mAbs could detect the CP4 EPSPS protein in the LM cotton traits MON1445 and MON88913 with high specificity, but not in any other LM cottons or non-LM cottons. These data indicate that these five mAbs to CP4 EPSPS could be successfully used for the further development of antibody-based detection methods to target CP4 EPSPS protein in LMOs.

• The Korean Journal of Mycology
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• v.33 no.2
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• pp.75-80
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• 2005

A carboxymethyl cellulase (CMCase) has been purified from Loweporus roseoalbus. The molecular weight of the purified CMCase was estimated to be 28.5 kDa by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis. The maximum activity of the purified CMCase was observed at pH 4.0 and $30^{\circ}C$, and stable for pH 3 to 5 to maintain 60% activity. The CMCase activity was activated by SDS and inhibited by PMSF and 1,10-phenanthroline. The enzyme activity was also decreased by the addition of ethylene diamine tetraacetic acid (EDTA), suggesting that the purified CMCase is metalloenzyme.

• Preventive Nutrition and Food Science
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• v.2 no.3
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• pp.250-254
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• 1997

The internal invertase of Rhodosporidium toruloids mating type a cells was purified to a single band on SDS-PAGE from cell-free extract by acid precipitation, ion exchange chromatogaphy andgel filtration. The determined molecular weight of he purified enzyme was about 95,000 by gel filtration and 100,000 daltons on SDS-polyacryamide gel electrophoresis. This enzyme didn't show any activity change by several metal ions except 15.4% decrease by {TEX}$Mn^{2+}${/TEX} and was strongly inhibited by 2-mercaptoethanol and SDS. The invertase maintained its activity at high level until 70℃, but inactivated at 80℃ almost completely. The optimal temperature and pH of the enzyme were about 60℃ and pH 5.0, respectively. The stable pH range of invertase was narrow from pH 3.0 to 6.0. The Km value and isoelectric point of enzyme were 3.4×{TEX}$10^{3}${/TEX} M, pH 4.4, respectively.

• Journal of Microbiology
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• v.35 no.4
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• pp.323-326
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• 1997

Toxin protein from Ustilago maydis virus SH14 isolated in Korea was purified using ethanol precipitation, cation exchange, gel filtration and anion exchange chromatography. The molecular weight of the purified protein was estimated to be 8.3 kDa by SDS-PAGE analysis. The Nterminal sequence of the protein is L-G-I-N-C(K)-R-G-S-S-Q--C(K)-G-L-S-G which is highly homologous with that of P4 toxin, but the amino acid composition and electrophoretic mobility in a native PAGE of the toxin protein were totally different from those of P4 toxin respectively. The SH14 toxin was shown to have immunological cross-reactivity about 50% with P4 toxin when examined by Western hybridization.

• Journal of Plant Biology
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• v.32 no.4
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• pp.331-341
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• 1989

DNA methyltransferase was purified 282.6-fold from Chlamydomonas reinhardtii 21gr (mt+) gametic cell to examine the effect of polyamine on the enzyme acctivity. Polyacrylamide gel electrophoresis(PAGE) revealed at least three bands(1 major band, 2 minor bands). Among these, the major band represents DNA methyltransferase. Polyacrylamide gel electrophoresis in the presence of 0.1% sodium dodecylsulfate(SDS-PAGE) revealed a major band with M.W. 60,000. DNA methyltransferase activity was inhibited more effectively by spermine than by spermidine, and the inhibition by putrescine was smaller than spermine and spermidine. DNA methyltransferase activity was inhibited by 40% and 53% at 5mM and 20mM spermine, respectively. In the case of spermidine, the inhibition was 35% at 10mM and 44% at 20mM. However, the inhibition by putrescine appeared only above 5mM and reached about 25% at 20mM.

• Bulletin of the Korean Chemical Society
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• v.25 no.5
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• pp.711-714
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• 2004

Peroxynitrite enters erythrocytes through band 3 anion exchanger and oxidizes cytosolic proteins therein. As a protein associated with band 3, carbonic anhydrase II may suffer from peroxynitrite-induced oxidative damages. Esterase activity of carbonic anhydrase II decreased as the concentration of peroxynitrite increased. Neither hydrogen peroxide nor hypochlorite affected the enzyme activity. Inactivation of the enzyme was in parallel with the release of zinc ion, which is a component of the enzyme's active site. SDS-PAGE of peroxynitrite-treated samples showed no indication of fragmentation but non-denaturing PAGE exhibited new bands with lower positive charges. Western analysis demonstrated that nitration of tyrosine residues increased with the peroxynitrite concentration but the sites of nitration could not be determined. Instead MALDI-TOF analysis identified tryptophan-245 as a site of nitration. Such modification of tryptophan residues is responsible for the decrease in tryptophan fluorescence. These results demonstrate that peroxynitrite nitrates tyrosine and tryptophan residues of carbonic anhydrase II without causing fragmentation or dimerization. The peroxynitrite-induced inactivation of the enzyme is primarily due to the release of zinc ion in the enzyme's active site.

• Microbiology and Biotechnology Letters
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• v.23 no.4
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• pp.411-415
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• 1995

The dextranase (EC 3.2.1.11) produced by Aspergillus ustus GR-98 was purified by the following sequential methods; salting-out and dialysis, gel filtration on BIO-GEL P-100, ion exchange chromatography on DEAH-cellulose, affinity chromatography on hydroxyapatite, and preparative electrophoresis. Three active fractions, dextranases 1, 11 and 111, were isolated in electrophoretically pure states, and specific activities of the dextranases were 1,276, 1,154 and 1,125 units/mg, the degrees of yield were 9.0, 3.6 and 2.2%, having 145, 131.1 and 127.8 times as those of culture filtrate in degree of purification, respectively. The enzyme purity was confirmed by the PAGE, SDS-PAGE and get permeation-HPLC.

• Preventive Nutrition and Food Science
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• v.7 no.1
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• pp.48-51
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• 2002

In order to study the reaction behavior of bovine holo- and apo-$\alpha$-lactalbumin ($\alpha$-La) during heat treatment at 65~10$0^{\circ}C$, the samples were analysed by first (ID)-and second-dimensional (2D) native-polyacrylamide gel electrophoresis (Native-PAGE) and sodium dodecylsulfate (SDS)-PAGE. When bolo-$\alpha$-La or apo- $\alpha$ -La were heated, they formed non-native, monomers, dimers and trimers. The apo-$\alpha$-La was more heat-sensitive than holo-$\alpha$-La. The monomers seemed to have the same composition as the native $\alpha$-La, but many of the disulfide bonds could be non-native.

• Journal of Microbiology and Biotechnology
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• v.22 no.1
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• pp.58-68
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• 2012

An investigation was conducted on the enhancement of production and purification of an oxidant and SDS-stable alkaline protease (BHAP) secreted by an alkalophilic Bacillus horikoshii, which was screened from the body fluid of a unique Korean polychaeta (Periserrula leucophryna) living in the tidal mud flats of Kwangwha Island in the Korean West Sea. A prominent effect on BHAP production was obtained by adding 2% maltose, 1% sodium citrate, 0.8% NaCl, and 0.6% sodium carbonate to the culturing medium. The optimal medium for BHAP production contained (g/l) SBM, 15; casein, 10; $K_2HPO_4$, 2; $KH_2PO_4$, 2; maltose, 20; sodium citrate, 10; $MgSO_4$, 0.06; NaCl, 8; and $Na_2CO_3$, 6. A protease yield of approximately 56,000 U/ml was achieved using the optimized medium, which is an increase of approximately 5.5-fold compared with the previous optimization (10,050 U/ml). The BHAP was homogenously purified 34-fold with an overall recovery of 34% and a specific activity of 223,090 U/mg protein using adsorption with Diaion HPA75, hydrophobic interaction chromatography (HIC) on Phenyl-Sepharose, and ion-exchange chromatography on a DEAE- and CM-Sepharose column. The purified BHAP was determined a homogeneous by SDS-PAGE, with an apparent molecular mass of 28 kDa, and it showed extreme stability towards organic solvents, SDS, and oxidizing agents. The $K_m$ and $k_{cat}$ values were 78.7 ${\mu}M$ and $217.4s^{-1}$ for N-succinyl-Ala-Ala-Pro-Phe-pNA at $37^{\circ}C$ and pH 9, respectively. The inhibition profile exhibited by PMSF suggested that the protease from B. horikoshii belongs to the family of serine proteases. The BHAP, which showed high stability against SDS and $H_2O_2$, has significance for industrial application, such as additives in detergent and feed industries.