• Title/Summary/Keyword: Functional characterization

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Cloning and Characterization of the pyrH Gene Encoding UMP-Kinase from Lactobacillus reuteri ATCC 55739

  • PARK JAE-YONG;NAM SU JIN;KIM JONG-HWAN;JEONG SEON-JU;KIM JUNG KON;HA YEONG LAE;KIM JEONG HWAN
    • Journal of Microbiology and Biotechnology
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    • v.15 no.3
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    • pp.525-531
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    • 2005
  • From a genomic library of Lactobacillus reuteri ATCC 55739, one clone, NE347, carrying a pyrH gene encoding UMP kinase, was identified. pNE347 carried a 1.88 kb EcoRI fragment and the pyrH was located in the middle of the insert. pyrH ORF was 723 bp in size and capable of encoding UMP kinase composed of 240 amino acid residues. tsf encoding an elongation factor-Ts and frr encoding a ribosomal recycling factor were present upstream and downstream of pyrH, respectively. When introduced into E. coli KUR1244, a pyrH-negative strain, pNE347 restored the ability to grow at $42^{\circ}C$, indicating that pyrH from L. reuteri synthesized functional UMP kinase in E. coli. Northern blot experiment showed that pyrH and frr were cotranscribed as a 1.4 kb single transcript. pyrH was overexpressed in E. coli by using a pET26b(+) vector, and a major 25 kDa protein band appeared on SDS-polyacrylamide gel.

Characterization of Mitochondrial Heat Shock Protein 75 (mtHSP75) of the Big-belly Seahorse Hippocampus abdominalis (빅벨리해마(Hippocampus abdominalis)에서의 Mitochondrial Heat Shock Protein 75 유전자의 특징과 발현 분석)

  • Ko, Jiyeon;Qiang, Wan;Lee, Sukkyoung;Bathige, S.D.N.K.;Oh, Minyoung;Lee, Jehee
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.48 no.3
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    • pp.354-361
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    • 2015
  • Mitochondrial heat shock protein 75 (mtHSP75) is a member of the HSP90 family and plays essential roles in refolding proteins of the mitochondrial matrix. Mitochondria provide energy in the form of ATP and generate reactive oxygen species (ROS). Heat shock proteins (HSPs) are activated in response to stress, and protect cells. In this study, we characterized the mtHSP75 of the big-belly seahorse Hippocampus abdominalis. The protein (BsmtHSP75) is encoded by an open reading frame (ORF) of 2,157 nucleotides, has 719 amino acids (aa), and is of molecular mass 82 kDa. BsmtHSP75 has two functional domains, a histidine kinase-like ATPase (HATPase_c) domain (123-276 aa) and an HSP90 family domain (302-718 aa). BsmtHSP75 was expressed in all tested tissues of healthy seahorses. The ovary contained the highest transcription level, followed (in order) by the blood, brain, and muscle. Pouch tissue showed the lowest expression level. The expression of BsmtHSP75 was significantly (P<0.05) up-regulated on viral or bacterial challenge, suggesting that BsmtHSP75 plays a role in the immune defense against bacterial and viral pathogens.

Improvement on the Antioxidant Activity of Instant Noodles Containing Enzymatic Extracts from Ecklonia cava and Its Quality Characterization (감태 효소 추출물을 이용한 즉석 국수의 항산화성 개선 및 품질 특성)

  • Heu, Min-Soo;Yoon, Min-Seok;Kim, Hyung-Jun;Park, Kwon-Hyun;Lee, Jong-Hyun;Jo, Mi-Ran;Lee, Jung-Suk;Jeon, You-Jin;Kim, Jin-Soo
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.43 no.5
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    • pp.391-399
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    • 2010
  • This study was conducted to improve the antioxidative activity of instant noodles containing enzymatic extracts from Ecklonia cava (EEC). EEC has relatively better antioxidative activity than extracts from other indigenous plants in Jeju Island. The EEC (2.5 mg/mL) had 82.5% for the hydroxy radical, 78.4% for the DPPH radical, and 64.9% for the superoxide anion radical scavenging activities, and 65.2% for the cell viability (100 ${\mu}g/mL$). According to the texture of the dough, the DPPH free radical scavenging of uncooked instant noodles, sensory evaluation of cooked instant noodles, and turbidity of the cooking drip, the optimal EEC concentration was 1.8% for the instant noodles. The major amino acids in the instant noodles with EEC were glutamic acid (24.2%), proline (10.2%), valine (10.0%), and isoleucine (12.3%). The zinc and iron in the instant noodles were enhanced by adding 1.5-1.8% EEC. The antioxidant activity of instant noodles with EEC was 75.4% for the hydroxy radical, 74.1% for the DPPH radical, and 51.2 % for the superoxide anion radical scavenging activities.

Characterization of Acid- and Pepsin-soluble Collagens from Rockfish Sebastes schlegeli Skin

  • Kim, Hyung-Jun;Jee, Seong-Joon;Yoon, Min-Suck;Youn, Mu-Ho;Kang, Kyung-Tae;Lee, Dong-Ho;Heu, Min-Soo;Kim, Jin-Soo
    • Fisheries and Aquatic Sciences
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    • v.12 no.1
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    • pp.6-15
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    • 2009
  • Biochemical and functional properties of acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) from rockfish skin were characterized. Yield of PSC (90.0%) was higher than that of ASC (63.2%). Both ASC and the PSC consisted of ${\alpha}1$ and ${\alpha}2$ chains, and $\alpha$-cross-linked components. According to the results of hydroxylation of proline and lysine, and FT-IR, no difference between the helical structure of ASC and PSC was identified. Thermal denaturation temperature (TDT) of ASC from rockfish skin was $22.8^{\circ}C$, the same as exhibited in PSC. Both ASC and PSC were higher in water absorption capacity (WAC) and oil absorption capacity (OAC) than other vegetable proteins. According to the results of emulsifying activity (EA) and cooking stability (CS), both ASC and PSC from rockfish skin were inferior compared to the commercial emulsifier (Tween-80). The results of FT-IR suggested that the structure of PSC was slightly different when compared to that of ASC. No differences in solubility were established between ASC and PSC from rockfish skin at various pH and NaCl concentrations.

Characterization of Chromatin Structure-associated Histone Modifications in Breast Cancer Cells

  • Hong, Chang-Pyo;Choe, Moon-Kyung;Roh, Tae-Young
    • Genomics & Informatics
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    • v.10 no.3
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    • pp.145-152
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    • 2012
  • Chromatin structure and dynamics that are influenced by epigenetic marks, such as histone modification and DNA methylation, play a crucial role in modulating gene transcription. To understand the relationship between histone modifications and regulatory elements in breast cancer cells, we compared our chromatin immunoprecipitation sequencing (ChIP-Seq) histone modification patterns for histone H3K4me1, H3K4me3, H3K9/16ac, and H3K27me3 in MCF-7 cells with publicly available formaldehyde-assisted isolation of regulatory elements (FAIRE)-chip signals in human chromosomes 8, 11, and 12, identified by a method called FAIRE. Active regulatory elements defined by FAIRE were highly associated with active histone modifications, like H3K4me3 and H3K9/16ac, especially near transcription start sites. The H3K9/16ac-enriched genes that overlapped with FAIRE signals (FAIRE-H3K9/14ac) were moderately correlated with gene expression levels. We also identified functional sequence motifs at H3K4me1-enriched FAIRE sites upstream of putative promoters, suggesting that regulatory elements could be associated with H3K4me1 to be regarded as distal regulatory elements. Our results might provide an insight into epigenetic regulatory mechanisms explaining the association of histone modifications with open chromatin structure in breast cancer cells.

Micro Porous Clay Mineral Absorption / Desorption Moisture-Proof Performance of The Atmospheric Humidity and Decomposing The Polyamide Adsorption Performance Characterization of Formaldehyde (미세 다공질 광물과 아미드계 분해제의 적용을 통한 건축자재의 습도 조절과 폼알데히드 분해 성능 특성 평가)

  • Lee, Che Cheol;Kim, Yun Hwan;Yun, Seng Hee
    • KIEAE Journal
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    • v.14 no.3
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    • pp.105-109
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    • 2014
  • The recent rising living standards, environment-friendly, well-being and health aspects of life in the basic gratification, as well as the desire for a pleasant environment emotionally environmentally friendly way of external space or industrial interest in the indoor environment, the manifestation. In particular, the biggest problem of the indoor environment has been emerged as a Sick House Syndrome indoor space that is provided to the building materials, and the impact on the domestic and the indoor environment, and clean the house in a health standards are specified as laws. The performance rating and the various materials to create environmentally-friendly standards for building materials. The more detail, Porous clay material, toxic substances released by applying the high humidity and the water itself, and to absorb the moisture, if the emissions, without a separate device, to maintain a comfortable indoor environment and at the same time, one of the causes of Sick House Syndrome breaking down harmful substances to absorb a comfortable indoor environment to maintain an environmentally-friendly building interior material studies. It is aimed at the development to multi-functional high performance eco-friendly construction materials, rather than through one feature performance, identify key features for national and international eco-friendly building materials can exert Water Vapour Adsorption raw, decomposed materials for the application and selection.

Synthesis and Characterization of New Polyaza Macrocyclic Nickel(Ⅱ) and Copper(Ⅱ) Complexes Two Nitrile or Imidate Ester Pendant Arms: Metal-Mediated Hydrolysis and Alcoholysis of the Nitrile Groups

  • Kang, Shin-Geol;Song, Jeong-Hoon;Jeong, Jong-Hwa
    • Bulletin of the Korean Chemical Society
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    • v.23 no.6
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    • pp.824-829
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    • 2002
  • New di-N-cyanomethylated tetraaza macrocycle 2.13-bis(cyanomethyl)-5.16-dimethyl-2,6,13,17-tetraazatricyclo[$16.4.0.0^7.12$]docosane $(L^2)$ has been prepared by the reaction of 3, 14-dimethyl-2,6,13,17-tetraazatricyclo $(L^1)$ with bromoacetonitrile. The square-planar complexes $[ML^2](ClO_4)_2(M=Ni(II)$ or Cu(II) can be prepared by the reaction of $L^2$ with the corresponding metal ion in acetonitrile. The cyanomethyl groups of $[ML^2](ClO_4)_2readily$ react with water to $yield[ML^3](ClO_4)_2$ containing pendant amide groups. The trans-octahedral complexes $[ML^4](ClO_4)_2$, in which two imidate ester groups are coordinated to the metal ion, can be also prepared by the reaction of $[ML^2](ClO_4)_2with$ methanol under mild conditions. The hydrolysis and alcoholysis reactions of $[ML^2](ClO_4)_2are$ promoted by the central metal ion, in spite of the fact that the cyanomethyl group is not involved in intramolecular coordination. The reactions are also promoted by a base such as triethylamine but are retarded by an $acid(HClO_4).Interestingly$, the imidate ester groups of $[ML^4]^2$ are unusually resistant to hydrolysis even in 0.1 M $HCIO_4$ or 0.1 M NaOH aqueous solution. Crystal structure of $[NiL^4](ClO_4)_2shows$ that the Ni-N (pendant imidate ester group) bond is rlatively strong; the Ni-N bond distance is shorter then the Ni-N(tertiary) distance and is similar to the Ni-N (secondary) distance.

Preparation and Characterization of Fluoroacrylate Copolymer by Emulsion Polymerization (유화 중합법에 의한 불소아크릴레이트 공중합체 제조)

  • 김동옥;김지현
    • Polymer(Korea)
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    • v.27 no.6
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    • pp.528-535
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    • 2003
  • The effects of surfactants, organic solvents, and functional monomers on the emulsion polymerization of perfluoroalkyleoylacryaltes and n-alkylacrylates were investigated. In particular, the dependence of the surface properties, contact angle and water repellency on the crystal melting temperature (T$\_$m/) of the fluorocopolymer and the variation of polymer latex particle sizes was investigated. Using WAXD experiments and synthesizing different types of fluorocopolymers which have fallowing fluoroacrylaytes [CH$_2$=CHCO$_2$CH,$_2$(CF$_2$CF$_2$) nH] (n = 4, 5 or 6), the relationship between the molecular packing structure of pendent side groups of fluorocopolymers and the surface properties was also investigated. We observed that the structure of primary carbon atoms of pendent side groups of fluorocopolymers plays key role in determining the surface properties.s.

Characterization of a Soil Metagenome-Derived Gene Encoding Wax Ester Synthase

  • Kim, Nam Hee;Park, Ji-Hye;Chung, Eunsook;So, Hyun-Ah;Lee, Myung Hwan;Kim, Jin-Cheol;Hwang, Eul Chul;Lee, Seon-Woo
    • Journal of Microbiology and Biotechnology
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    • v.26 no.2
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    • pp.248-254
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    • 2016
  • A soil metagenome contains the genomes of all microbes included in a soil sample, including those that cannot be cultured. In this study, soil metagenome libraries were searched for microbial genes exhibiting lipolytic activity and those involved in potential lipid metabolism that could yield valuable products in microorganisms. One of the subclones derived from the original fosmid clone, pELP120, was selected for further analysis. A subclone spanning a 3.3 kb DNA fragment was found to encode for lipase/esterase and contained an additional partial open reading frame encoding a wax ester synthase (WES) motif. Consequently, both pELP120 and the full length of the gene potentially encoding WES were sequenced. To determine if the wes gene encoded a functioning WES protein that produced wax esters, gas chromatography-mass spectroscopy was conducted using ethyl acetate extract from an Escherichia coli strain that expressed the wes gene and was grown with hexadecanol. The ethyl acetate extract from this E. coli strain did indeed produce wax ester compounds of various carbon-chain lengths. DNA sequence analysis of the full-length gene revealed that the gene cluster may be derived from a member of Proteobacteria, whereas the clone does not contain any clear phylogenetic markers. These results suggest that the wes gene discovered in this study encodes a functional protein in E. coli and produces wax esters through a heterologous expression system.

Molecular Characterization of Cold-Inducible ${\beta}$-Galactosidase from Arthrobacter sp. ON14 Isolated from Antarctica

  • Xu, Ke;Tang, Xixiang;Gai, Yingbao;Mehmood, Muhammad Aamer;Xiao, Xiang;Wang, Fengping
    • Journal of Microbiology and Biotechnology
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    • v.21 no.3
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    • pp.236-242
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    • 2011
  • A psychrotrophic bacterium, Arthrobacter sp. ON14, isolated from Antarctica, was shown to exhibit a high ${\beta}$-galactosidase activity at a low temperature. A genomic library of ON14 was constructed and screened for ${\beta}$-galactosidase genes on functional plates containing 5-bromo-4-chloro-3-indolyl-${\beta}$-D-galactopyranoside (X-gal) as the substrate. Two different ${\beta}$-galactosidase genes, named as galA, galB, were found in ON14. Computational analyses of the genes revealed that the encoded protein GalA belongs to family 2 of glycosyl hydrolysases and is a cold-active protein, whereas GalB belongs to family 42 of glycosyl hydrolysases and is a mesophilic protein. Reverse transcription analyses revealed that the expression of galA is highly induced at a low temperature ($4^{\circ}C$ ) and repressed at a high temperature ($28^{\circ}C$ ) when lactose is used as the sole carbon source. Conversely, the expression of galB is inhibited at a low temperature and induced at a high temperature. The purified GalA showed its peak activity at $15^{\circ}C$ and pH 8. The mineral ions $Na^+$, $K^+$, $Mg^{2+}$, and $Mn^{2+}$ were identified as enzyme activators, whereas $Ca^{2+}$ had no influence on the enzyme activity. An enzyme stability assay revealed that the activity of GalA is significantly decreased when it is incubated at $45^{\circ}C$ for 2 h, and all its activity is lost when it is incubated at $50^{\circ}C$.