• Journal of Marine Bioscience and Biotechnology
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• v.1 no.4
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• pp.268-274
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• 2006

Bacillus sp. DYL130 producing biosurfactant was isolated from soil samples in the Duck-yu mountain and identified as Bacillus sp. by analysis of 16S rDNA sequence. Purification of the biosurfactant was performed by using affinity chromatography and TLC. The biosurfactant of culture medium from Bacillus sp. DYL130 was eluted with 100% methanol using affinity chromatography. To remove methanol, a rotary evaporator was used and enrichment sample was dissolved in alkaline water(pH 10). The purified biosurfactant was identified by TLC. It was confirmed that the Rf value of the biosurfactant was 0.78. Antifungal activity against Botrytis cineria was showed the strongly activity as active antagonist. Maximum emulsification activity and stability were obtained from soybean oil. The critical micelle concentration (CMC) of purified biosurfactant was 35mg/l and the purified biosurfactant inhibited biofilm forming by Bacillus sp..

• Journal of Life Science
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• v.27 no.11
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• pp.1233-1244
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• 2017

CDPKs have pivotal roles in plant $Ca^{2+}$-mediated transduction signaling. A total of 29 CDPK genes have been identified in rice (Oryza sativa L.), but their key functions have not been completely noted. This study focused on the OsCPK11 gene, which has not been studied, to determine its functional characteristics. A study of tissue-specific expressions revealed that the OsCPK11 gene is expressed in young leaves, mature leaves and flowers of rice. An expression of the gene was also confirmed in gibberellin-treated aleurone layers of rice. Regarding the phenotypic characteristics of Tos17-inserted OsCPK11 mutants, the heights of the mutants were not distinguishable from the heights of wild type plants, but the number of caryopses and the caryopses' weights were significantly statistically different. In addition, many grains of the mutants had white belly materials in their endosperm. The cDNA of the OsCPK11 was cloned, and an OsCPK11 protein of about 60.5 kD was obtained by using a GST affinity chromatography and an SDS-PAGE. An analysis of the amino-acid sequence of the protein indicated that the OsCPK11 protein has the structural characteristics of typical CDPKs. The results provided useful information about the functions of the OsCPK11 gene and further noted the roles CDPKs have in $Ca^{2+}$-mediated signaling in plants.

• The Korean Journal of Mycology
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• v.32 no.2
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• pp.112-118
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• 2004

Laccase produced by Trametes hirsuta S1 isolated from Korea was partially purified and characterized using ultrafiltration, anion exchange chromatography and affinity chromatography. The laccase was produced as the predominant extracellular enzyme during primary metabolism. Neither lignin peroxidase nor veratryl alcohol oxidase (VAO) were detected in the culture fluid. Addition of 2,5-xylidine enhanced 4-fold laccase production. Purified laccase was a single polypeptide having a molecular mass of approximately 66 kDa, as determined by SDS-polyacrylamide gel electrophoresis, and carbohydrate content of 12%. $K_{m}\;and\;V_{max}$ values for laccase with ABTS [2,2-azinobis (3-ethylbenzthiazoline 6-sulfonic acid)] as a substrate (Lineweaver-Burk plot) was determined to $51.2\;{\mu}M\;and\;56.8\;{\mu}mole$, respectively. The optimal pH for laccase activity was found to be 3.0. The enzyme was very stable for 1 hour at $50^{\circ}C$. Half-life ($t_{1/2}$) of the enzyme was about 20 min at $70^{\circ}C$. Spectroscopic analysis of purified enzyme indicated that the enzyme was typical of copper-containing protein. Substrate specificity and inhibitor studies for laccase also indicated to be a typical fungal laccase. The N-terminal amino acid sequence of the T. hirsuta S1 laccase showed 100% of homology to those of laccase from C. hirsutus.

• Korean Journal of Food Science and Technology
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• v.34 no.4
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• pp.552-558
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• 2002

Polyphenol oxidase from Flammulina velutipes was purified and characterized. Purification of polyphenol oxidase was achieved by ammonium sulfate precipitation, Superdex G-200 gel filtration chromatography, Phenyl superose affinity chromatography, Mono-Q anion exchange chromatography and Superdex S-200 gel filtration chromatography on FPLC. After these purification steps specific activity of purified polyphenol oxidase increased to 199.1 units/mg. Polyphenol oxidase from F. velutipes was composed of a single polypeptide with molecular weight of about 40 kDa. Optimum pH and temperature for the enzyme reaction were found to be 6.0 and $25^{\circ}C$, respectively. The activity of the enzyme gradually decreased at acidic pH between 3 and 5, and the enzyme lost its activity at alkaline pH between 8 and 10. This enzyme exhibited high substrate specificity to o-diphenols. Km-values for L-DOPA and caffeic acid were found to be 3.97 mM and 1.78 mM, respectively. 2-mercaptoethanol, L-ascorbic acid, sodium bisulfite, EDTA and $Mg^{2+}$ inhibited the activity of pholyphenol oxidase and $Cu^{2+}$, $Fe^{2+}$, $Zn^{2+}$ and $Ni^{2+}$ increased enzyme activity. The activity of enzyme was well maintained at $-70^{\circ}C$ for over 4 months, and at $-20^{\circ}C$ for 1 months.

• Journal of Dairy Science and Biotechnology
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• v.15 no.1
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• pp.33-44
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• 1997

We investigated the immunological function of cheese whey protein concentrate (CWPC), which is a by-product of cheese production, using mitogenic activity in murine splenocytes as an index. A fraction isolated by gel filtration and anion exchange chromatography of CWPC showed high mitogenic activity, comparable to the activity of lipopolysaccharide (LPS). The fraction was detected as a single band on SDS-PAGE. It contained calcium, inorganic phosphorus, and carbo-hydrate, indicating the active component to be a glycophosphopeptide (GPP) Since pronase digestion of GPP did not reduce its mitogenic activity, carbohydrate rather than peptide may be important in the activity, When applied on an anti-${\beta}$-caseinophosphopeptide (${\beta}$-CPP ) antibody affinity column, the GPP was separated into two components, one with affinity to ${\beta}$-CPP and the other without such affinity. Both the components contained N-linked oligosaccharide chains and had the mitogenic activity. These results demonstrate that cheese whey contains a GPP having strong mitogenic activity

• Korean Journal of Food Science and Technology
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• v.32 no.5
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• pp.1079-1086
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• 2000

Pectinesterase inhibitor(PEI) of ripened kiwi fruit(Actinidia chinensis) was separated with affinity chromatography using CNBr-activated Sepharose 4B being covalently bound by orange pectinesterse(PE). The affinity resin strongly and selectively bound PEI, which could be eluted in high yield as a single peak by pH 9.5 without loss of inhibitory activity. The separated PEI had maintained almost inhibitory activity at $-25^{\circ}C$ and $5^{\circ}C$ during 30 days but lost it at room temperature in 4 weeks. The PEI possessed a molecular weight of 16.6 KDa, as estimated by 12.5% SDS-PAGE. PEI had optimum pH of 7.5, optimum temperature of below $10^{\circ}C$ and stability up to $70^{\circ}C$. Also, optimum inhibitory activity for PEI was obtained in 0.2 M NaCl of substrate solutions. The kind of inhibition on tomato pectinesterase was found to be noncompetitive, using citrus pectin as substrate. Fresh orange juice added with crude PEI extracts maintained almost the same cloud stability as pasteurized juice. In case of apple juice, the addition of crude PEI extracts to apple juice had decrease of L-ascorbic acid with nearly no effect on cloud loss.

• Journal of Ginseng Research
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• v.46 no.6
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• pp.750-758
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• 2022

Background: Mild cognitive impairment (MCI) is a transitional condition between normality and dementia. Ginseng is known to have effects on attenuating cognitive deficits in neurogenerative diseases. Ginsenosides are the main bioactive component of ginseng, and their protein targets have not been fully understood. Furthermore, no thorough analysis is reported in ginsenoside-related protein targets in MCI. Methods: The candidate protein targets of ginsenosides in brain tissues were identified by drug affinity responsive target stability (DARTS) coupled with label-free liquid chromatography-mass spectrometry (LC-MS) analysis. Network pharmacology approach was used to collect the therapeutic targets for MCI. Based on the above-mentioned overlapping targets, we built up a proteineprotein interaction (PPI) network in STRING database and conducted gene ontology (GO) enrichment analysis. Finally, we assessed the effects of ginseng total saponins (GTS) and different ginsenosides on mitochondrial function by measuring the activity of the mitochondrial respiratory chain complex and performing molecular docking. Results: We screened 2526 MCI-related protein targets by databases and 349 ginsenoside-related protein targets by DARTS. On the basis of these 81 overlapping genes, enrichment analysis showed the mitochondria played an important role in GTS-mediated MCI pharmacological process. Mitochondrial function analysis showed GTS, protopanaxatriol (PPT), and Rd increased the activities of complex I in a dose-dependent manner. Molecular docking also predicted the docking pockets between PPT or Rd and mitochondrial respiratory chain complex I. Conclusion: This study indicated that ginsenosides might alleviate MCI by targeting respiratory chain complex I and regulating mitochondrial function, supporting ginseng's therapeutic application in cognitive deficits.

• Bulletin of the Korean Chemical Society
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• v.33 no.10
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• pp.3248-3252
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• 2012

Vibrio extracellular metalloprotease (vEP), secreted from Vibrio vulnificus, shows various proteolytic function such as prothrombin activation and fibrinolytic activities. Premature form of vEP has an N-terminal (nPP) and a C-terminal (C-ter100) region. The nPP and C-ter100 regions are autocleaved for the matured metalloprotease activity. It has been proposed that two regions play a key role in regulating enzymatic activity of vEP. Especially, C-ter100 has a regulatory function on proteolytic activity of vEP. C-ter100 domain has been cloned into the E. coli expression vectors, pET32a and pGEX 4T-1 with TEV protease cleavage site and purified using gel-filtration chromatography followed by affinity chromatography. To understand how C-ter100 modulates proteolytic activity of vEP, structural studies were performed by heteronuclar multi-dimensional NMR spectroscopy. Backbone $^1H$, $^{15}N$ and $^{13}C$ resonances were assigned by data from standard triple resonance and HCCH-TOCSY experiments. The secondary structures of vEP C-ter100 were determined by TALOS+ and CSI software based on hydrogen/deuterium exchange. NMR data show that C-ter100 of vEP forms a ${\beta}$-barrel structure consisting of eight ${\beta}$-strands.

• Korean Journal of Microbiology
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• v.34 no.1_2
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• pp.43-50
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• 1998

Induction and purification of the GroEL homolog from Streptococcus pneumolliae were characterized. The stress conditions were determined by temperature, ethanol, NaCI, $H_2O_2$ methyl methanesulfonate, and ethyl methanesulfonate. And stress induced proteins were analyzed using [$^{35}S$]-methionine labeling method. Heat shock induced the synthesis of a set of about 3 heat shock proteins (hsps) (65, 73, and 84-kDa). Of those 3 hsps, a 65 kDa protein, hsp65, was purified by DEAE-Sephacel and ATP-agarose column chromatography, and used for antibody preparation. Immunoblot analysis employing antisera raised against pneumococcus hsp65 demonstrated cross-reactivity with a 60 kDa protein in Eschericha coli. Also cross reaction of the purified p65 with anti-Escherichia coli GroEL monoclonal antibody demonstrated that pneumococcal hsp65 is the GroEL homolog.

• Korean Journal of Microbiology
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• v.24 no.3
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• pp.251-262
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• 1986

High-molecular-weight ${\beta}-glucosidase$ (EC 3.2.1.21) was purified from the culture filtrate of Trichoderma koningii through a four-step procedure including chromatography on Bio-Gel P-150, DEAE-Sephadex A-50 and SP-Sephadex C-50; and chromatofocusing on Polybuffer exchanger PBE 94. The molecular weight of the enzyme was determined to be about 101,000 by SDS-polyacrylamide gel electrophoreses, and the isoelectric point was estimated to be 4.96 by analytical isoelectric focusing. The temperature optimum for activity was about $55^{\circ}C$, and the pH optimumwas 3.5. The enzyme was considerably thermostable, for no loss of activity was observed when the enzyme was preincubated at $60^{\circ}C$ for 5h. Km values for cellobiose, gentiobiose, sophorose, salicin and $p-nitrophenyl-{\betha}-D-glucoside$ were 99.2, 14.7, 7.09, 3.15 and 0.70 mM, respectively, which indicates that the enzyme has much higher affinity towards $p-nitrophenyl-{\betha}-D-glucoside$ than towards the other substrates, especially cellobiose. Substrate inhibition by $p-nitrophenyl-{\betha}-D-glucoside$ and salicin was observed at the conecntrations exceeding 5mM. Gluconolactone was a powerful inhibitor against the action of the enzyme on $p-nitrophenyl-{\betha}-D-glucoside\;(K_i\;37.9\;{\mu}M)$, wherease glucose was much less effective ($K_i$ 1.95 mM). Inhibition was of the competitive type in each case. Transglucosylation activity was detected shen the readtion products formed from $p-nitrophenyl-{\betha}-D-glucoside$ by the enzyme were analysed using high-performance liquid chromatography.