• 제목/요약/키워드: metabolic characterization

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Characterization and refinement of enzyme of the gene encoding catechol 1,2-dioxygenase from Phenol-degrading, Rhodococcus sp.

  • 이희정;박근태;박재림;이상준
    • Proceedings of the Korean Environmental Sciences Society Conference
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    • 2002.05b
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    • pp.209-212
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    • 2002
  • The heavy use of petroleum products in modern livings has brought ubiquitous environmental contaminants of aromatic compounds, which persist in aquatic and geo-environment without the substantial degradation. The persistence and accumulation of the aromatic compounds, which include xylene, phenol, toluene, phthalate, and so on are known to cause serious problems in our environments. Some of soil and aquatic microorganisms facilitate their growth by degrading aromatic compounds and utilizing degrading products as growth substrates, the biodegradation helps the reentry of carbons of aromatic compounds, preventing their accumulation in our environments. The metabolic studies on the degradation of aromatic compounds by microoganlsms were extensively carried out along with their genetic studies. A Rhodococcus sp. isolated in activated sludges has shown the excellent ability to grow on phenol as a sole carbon source. In the present study investigated a gene encoding phenol-degrading enzymes from a Rhodococcus sp.

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Kinetic Properties of Extracted Lactate Dehydrogenase and Creatine Kinase from Mouse Embryonic Stem Cell- and Neonatal-derived Cardiomyocytes

  • Zonouzi, Roseata;Ashtiani, Saeid Kazemi;Hosseinkhani, Saman;Baharvand, Hossein
    • BMB Reports
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    • v.39 no.4
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    • pp.426-431
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    • 2006
  • Embryonic stem cells (ESCs), representing a population of undifferentiated pluripotent cells with both self-renewal and multilineage differentiation characteristics, are capable of spontaneous differentiation into cardiomyocytes. The present study sought to define the kinetic characterization of lactate dehydrogenase (LDH) and creatine kinase (CK) of ESC- and neonatal-derived cardiomyocytes. Spontaneously differentiated cardiomyocytes from embryoid bodies (EBs) derived from mouse ESC line (Royan B1) and neonatal cardiomyocytes were dispersed in a buffer solution. Enzymes were extracted by sonication and centrifugation for kinetic evaluation of LDH and CK with spectrophotometric methods. While a comparison between the kinetic properties of the LDH and CK of both groups revealed not only different Michaelis constants and optimum temperatures for LDH but also different Michaelis constants and optimum pH for CK, the pH profile of LDH and optimum temperature of CK were similar. In defining some kinetic properties of cardiac metabolic enzymes of ESC-derived cardiomyocytes, our results are expected to further facilitate the use of ESCs as an experimental model.

Characterization of Spermidine Transport System in a Cyanobacterium, Synechocystis sp. PCC 6803

  • Raksajit, Wuttinun;Yodsang, Panutda;Maenpaa, Pirkko;Incharoensakdi, Aran
    • Journal of Microbiology and Biotechnology
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    • v.19 no.5
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    • pp.447-454
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    • 2009
  • The transport of spermidine into a cyanobacterium, Synechocystis sp. pec 6803, was characterized by measuring the uptake of $^{14}C$-spermidine. Spermidine transport was shown to be saturable with an apparent affinity constant ($K_m$) value of $67{\mu}M$ and a maximal velocity ($V_{max}$) value of 0.45 nmol/min/mg protein. Spermidine uptake was pH-dependent with the pH optimum being 8.0. The competition experiment showed strong inhibition of spermidine uptake by putrescine and spermine, whereas amino acids were hardly inhibitory. The inhibition kinetics of spermidine transport by putrescine and spermine was found to be noncompetitive with $K_i$ values of 292 and $432{\mu}M$, respectively. The inhibition of spermidine transport by various metabolic inhibitors and ionophores suggests that spermidine uptake is energy-dependent. The diminution of cell growth was observed in cells grown at a high concentration of NaCl. Addition of a low concentration of spermidine at 0.5 mM relieved growth inhibition by salt stress. Upshift of the external osmolality generated by either NaCl or sorbitol caused an increased spermidine transport with about 30-40% increase at 10 mosmol/kg upshift.

Characterization of Physiological Changes in $S3H5/\gamma{2bA2}$ Hybridoma Cells During Adaptation to Low Serum Media

  • Lee, Gyun-Min;Joanne, Savinell
    • Journal of Microbiology and Biotechnology
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    • v.2 no.2
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    • pp.141-151
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    • 1992
  • Physiological changes of the murine hybridoma cell line $S3H5/\gamma{2bA2}$ during adaptation to RPMI 1640 medium with 1%(v/v) fetal bovine serum were characterized in terms of cell growth, antibody production, morphology, and metabolic quotients. Cells adapted to 1% serum medium in T-flasks became sensitive to shear induced by mechanical agitation and required at least 5% serum in the medium or spent medium for cell growth in spinner flasks, while cells adapted to 10% serum medium in T-flasks could grow in 1% serum medium in spinner flasks. Consequently, long-term adaptation to low serum media may not give the expected growth enhancement. After adaptation to 1% serum medium, changes in cell morphology were observed. The cells in 10% serum medium were uniform and circular, while cells in 1% medium were irregularly shaped. The DNA contents, which were measured by flow cytometry, were almost constant among the cells in the range of 1% to 10%. Further, no significant changes in energy metabolism and specific monoclonal antibody production rate were observed among these cells.

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Crystal Structure and Biochemical Characterization of Xylose Isomerase from Piromyces sp. E2

  • Son, Hyeoncheol Francis;Lee, Sun-Mi;Kim, Kyung-Jin
    • Journal of Microbiology and Biotechnology
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    • v.28 no.4
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    • pp.571-578
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    • 2018
  • Biofuel production using lignocellulosic biomass is gaining attention because it can be substituted for fossil fuels without competing with edible resources. However, because Saccharomyces cerevisiae does not have a ${\text\tiny{D}}$-xylose metabolic pathway, oxidoreductase or isomerase pathways must be introduced to utilize ${\text\tiny{D}}$-xylose from lignocellulosic biomass in S. cerevisiae. To elucidate the biochemical properties of xylose isomerase (XI) from Piromyces sp. E2 (PsXI), we determine its crystal structure in complex with substrate mimic glycerol. An amino-acid sequence comparison with other reported XIs and relative activity measurements using five kinds of divalent metal ions confirmed that PsXI belongs to class II XIs. Moreover kinetic analysis of PsXI was also performed using $Mn^{2+}$, the preferred divalent metal ion for PsXI. In addition, the substrate-binding mode of PsXI could be predicted with the substrate mimic glycerol bound to the active site. These studies may provide structural information to enhance ${\text\tiny{D}}$-xylose utilization for biofuel production.

Characterization of biphenyl biodegradation, and regulation of iphenyl catabolism in alcaligenes xylosoxydans

  • Lee, Na-Ri;On, Hwa-Young;Jeong, Min-Seong;Kim, Chi-Kyung;Park, Yong-Keun;Ka, Jong-Ok;Min, Kyung-Hee
    • Journal of Microbiology
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    • v.35 no.2
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    • pp.141-148
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    • 1997
  • Alcaligenes xylosoxydans strain SMN3 capable of utilizing biphenyl grew not only on phenol, and benzoate, but also on salicylate. Catabolisms of biphenyl and salicylate appear to be interrelated since benzoate is a common metabolic intermediate of these compounds. Enzyme levels in the excatechol 2. 3-dioxygenas which is meta-cleavage enzyme of catechol, but did not induce catechol 1, 2-dioxygenase. All the oxidative enzymes of biphenyl and 2, 3,-dihydroxybiphenyl (23DHBP) were induced when the cells were grown on biphenyl and salicylate, respectively. Biphenyl and salicylate could be a good inducer in the oxidation of biphenyl and 2, 3-dihydroxybiphenyl. The two enzymes for the degradation of biphenyl and salicylate were induced after growth on either biphenyl or salicylate, suggesting the presence of a common regulatory element. However, benzoate could not induce the enzymes responsible for the oxidation of these compounds. Biphenyl and salicylate were good inducers for indigo formation due to the activity of biphenyl dioxygenase. These results suggested that indole oxidation is a property of bacterial dioxygenase that form cis-dihydrodiols from aromatic hydrocarbon including biphenyl.

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Daily Profiles of Blood Insulin, Insulin-Like Growth Factor-I, Thyroxine and Triiodothyronine in Ewes Under Three Levels of Feed Intake

  • Caldeira, R.M.;Vasques, M.I.;Vaz Portugal, A.
    • Asian-Australasian Journal of Animal Sciences
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    • v.13 no.8
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    • pp.1121-1126
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    • 2000
  • Daily variation in the serum concentrations of insulin and insulin-like growth factor-I and in the plasma concentrations of thyroxine and triiodothyronine were evaluated in ewes fed 30%, 100% and 200% of theoretical maintenance energy requirements. The single daily meal has had significant effects (p<0.05) on almost all profiles. In general, serum or plasma hormone concentrations have increased after the meal, in particular at the two higher levels of energy intake. In the group submitted to the lowest level of energy intake, the consequences of the meal on circulating levels were almost imperceptible. The effects of feeding levels on serum or plasma concentrations have widely varied among hormones, not showing any objective pattern or relationship. Because these variations may affect the interpretation of these blood indicators, knowledge of daily profiles and of the effect of feed level must be considered. In order to maximize the diagnostic value of those indicators, the most suitable times for blood collection seem to be 16 h after the meal and (or) just before the meal. The collection 16 h after the meal apparently allows the characterization of a relatively steady metabolic state, intermediate between the close effects of food intake and the final phase of the intensification of body reserves mobilization. The collection just before the meal will give a good indication of the level of activity of those mobilization mechanisms.

Expression and Characterization of Truncated Recombinant Human Cytochrome P450 2J2

  • Park, Hyoung-Goo;Lim, Young-Ran;Han, Songhee;Kim, Donghak
    • Toxicological Research
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    • v.30 no.1
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    • pp.33-38
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    • 2014
  • The human cytochrome P450 2J2 catalyzes an epoxygenase reaction to oxidize various fatty acids including arachidonic acid. In this study, three recombinant enzyme constructs of P450 2J2 were heterologously expressed in Escherichia coli and their P450 proteins were successfully purified using a $Ni^{2+}$-NTA affinity column. Deletion of 34 amino acid residues in N-terminus of P450 2J2 enzyme (2J2-D) produced the soluble enzyme located in the cytosolic fraction. The enzymatic analysis of this truncated protein indicated the typical spectral characteristics and functional properties of P450 2J2 enzyme. P450 2J2-D enzymes from soluble fraction catalyzed the oxidation reaction of terfenadine to the hydroxylated product. However, P450 2J2-D enzymes from membrane fraction did not support the P450 oxidation reaction although it displayed the characteristic CO-binding spectrum of P450. Our finding of these features in the N-terminal modified P450 2J2 enzyme could help understand the biological functions and the metabolic roles of P450 2J2 enzyme and make the crystallographic analysis of the P450 2J2 structure feasible for future studies.

Eight unrecorded bacterial species isolated from soil and marine sediment in Korea

  • Kim, Minji;Lee, Ki-Eun;Cha, In-Tae;Lee, Byoung-Hee;Park, Soo-Je
    • Journal of Species Research
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    • v.9 no.4
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    • pp.339-345
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    • 2020
  • The Earth contains billions of microbial species, although the vast majority cannot be cultured in laboratories and are thus considered unidentified and uncharacterized. Extremophiles are microorganisms that thrive in extreme conditions, including temperature, salinity, and pH. Extremophilic microorganisms have provided important insights for biological, metabolic, and evolutionary studies. Between 2017 and 2019, as part of a comprehensive investigation to identify bacterial species in Korea, eight bacterial strains were isolated from marine and non-marine environments in Jeju Island. These strains were cultured under extreme salinity or pH conditions. Phylogenetic analysis using 16S ribosomal RNA(rRNA) gene sequencing indicated that all eight strains belonged to the phyla Gammaproteobacteria, Bacilli, and Alphaproteobacteria. Based on their high 16S rRNA gene sequence similarities(>98.7%) and the formation of strong monophyletic clades with their closest related species, all isolated strains were considered as an unrecorded strain, previously unidentified species. Gram stain reaction, culture conditions, colony and cell morphology, biochemical characteristics, isolation source, and National Institute of Biological Resources(NIBR) IDs are described in this article. The characterization of these unrecorded strains provides information on microorganisms living in Korea.

Isolation and Characterization of Malate Dehydrogenase Gene from Panax ginseng C.A. Meyer (고려인삼에서 Malate Dehydrogenase 유전자의 분리 및 분석)

  • Kim, Yu-Jin;Shim, Ju-Sun;Lee, Jung-Hye;Jung, Dae-Young;In, Jun-Gyo;Lee, Bum-Soo;Min, Byung-Hoon;Yang, Deok-Chun
    • Korean Journal of Medicinal Crop Science
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    • v.16 no.4
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    • pp.261-267
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    • 2008
  • Malate dehydrogenase is a ubiquitous enzyme in plants, involving in a range of metabolic processes depending on its subcellular location. A malate dehydrogenase (PgMDH) cDNA was isolated and characterized from the root of Panax ginseng C. A. Meyer. The deduced amino acid sequence of PgMDH showed high similarity with the NAD-dependent mitochondrial malate dehydrogenase from Glycinemax (P17783), Eucalyptus gunnii (P46487), and Lycopersicon esculentum (AAU29198). And the segment of a malate dehydrogenase gene was amplified through RT-PCR. The expression of PgMDH was increased after treatments of chilling, salt, UV, cadmium or copper treatment.