Characterization of glycosyl hydrolase family 3 β-N-acetylglucosaminidases from Thermotoga maritima and Thermotoga neapolitana

Choi, K.H.,Seo, J.Y.,Park, K.M.,Park, C.S.,Cha, J. Society for Bioscience and Bioengineering, Japan ; 2009 Journal of bioscience and bioengineering Vol.108 No.6

The genes encoding β-N-acetylglucosaminidase (nagA and cbsA) from Thermotoga maritima and Thermotoga neapolitana were cloned and expressed in Escherichia coli in order to investigate whether Thermotoga sp. is capable of utilizing chitin as a carbon source. NagA and CbsA were purified to homogeneity by HiTrap Q HP and Sephacryl S-200 HR column chromatography. Both enzymes were homodimers containing a family 3 glycoside hydrolase (GH3) catalytic domain, with a monomer molecular mass of 54 kDa. The optimal temperatures and pHs for the activities of the β-N-acetylglucosaminidases were found to be 65-75 <SUP>o</SUP>C and 7.0-8.0, respectively. Both enzymes hydrolyzed chitooligomers such as di-N-acetylchitobiose and tri-N-acetylchitotriose, and synthetic substrates such as p-nitrophenyl-β-d-glucose (pNPGlc), p-nitrophenyl N-acetyl β-d-glucosamine (pNPGlcNAc), p-nitrophenyl di-N-acetyl β-d-chitobiose (pNPGlcNAc<SUB>2</SUB>) and p-nitrophenyl tri-N-acetyl β-d-chitotriose (pNPGlcNAc<SUB>3</SUB>). However, the enzymes had no activity against p-nitrophenyl-β-d-galactose (pNPGal) and p-nitrophenyl N-acetyl β-d-galactosamine (pNPGalNAc) or highly polymerized chitin. The k<SUB>cat</SUB> and K<SUB>m</SUB> values were determined for pNPGlcNAc, pNPGlcNAc<SUB>2</SUB> and pNPGlcNAc<SUB>3</SUB>. The k<SUB>cat</SUB>/K<SUB>m</SUB> value for pNPGlcNAc was the highest among three synthetic substrates. NagA and CbsA initially hydrolyzed p-nitrophenyl substrates to give GlcNAc, suggesting that the enzymes have exo-activity with chitin oligosaccharides from the non-reducing ends, like other β-N-acetylglucosaminidases. However, NagA and CbsA can be distinguished from other GH3-type β-N-acetylglucosaminidases in that they are highly active against di-N-acetylchitobiose. Thus, the present results suggest that the physiological role of both enzymes is to degrade the chitooligosaccharides transported through membrane following hydrolysis of chitin into β-N-acetylglucosamine to be further metabolized in Thermotoga sp.

Improved production of phleichrome from the phytopathogenic fungus Cladosporium phlei using synthetic inducers and photodynamic ROS production by phleichrome

So, K.K.,Jo, I.S.,Chae, M.S.,Kim, J.M.,Chung, H.J.,Yang, M.S.,Kim, B.T.,Kim, J.K.,Choi, J.K.,Kim, D.H. Society for Bioscience and Bioengineering, Japan ; 2015 Journal of bioscience and bioengineering Vol.119 No.3

Two different diketopiperazines, cyclo-(l-Pro-l-Leu) and cyclo-(l-Pro-l-Phe), which were isolated from the culture filtrate of Epichloe typhina and found to be inducers of phleichrome production, were chemically synthesized and evaluated for use in the improved production of phleichrome from wild-type and UV-mutagenized strains (M0035) of Cladosporium phlei. When supplemented with PDA and V8 juice agar media, both inducers showed significant increases in the production of phleichrome. Phleichrome production was increased in a dose-dependent manner up to a concentration of maximum yield for both inducers. No further significant induction was observed by supplementing inducers over the concentration of maximum yield. Among the two inducers, cyclo-(l-Pro-l-Phe) showed better inducing capability than cyclo-(l-Pro-l-Leu). The maximum yield was observed from the M0035 strain grown on V8 juice media supplemented with 150 μM cyclo-(l-Pro-l-Phe), which was estimated to be 232.6 mg of phleichrome per gram of mycelia and 10.2 mg of secreted phleichrome per 20 agar-plugs. Interestingly, growth inhibition was observed on V8 juice agar media with 100, 150, and 200 μM cyclo-(l-Pro-l-Phe) but not on PDA with the same amount of inducer, which suggests that the inhibitory effect might be through the overproduction of phleichrome rather than the toxic effect of the inducer itself. Superoxide production by purified phleichrome was dramatically stimulated upon illumination, thus demonstrating photodynamic production of superoxide in vitro by phleichrome.

Synthesis and Thermoelectric Properties of Ce1−z Pr z Fe4−x Co x Sb12 Skutterudites

Song, K. M.,Shin, D. K.,Jang, K. W.,Choi, S. M.,Lee, S.,Seo, W. S.,Kim, I. H. Springer Science + Business Media 2017 Journal of electronic materials Vol.46 No.5

<P>p-Type Ce1-z Pr (z) Fe4-x Co (x) Sb-12 skutterudites were prepared by encapsulated melting, quenching, annealing, and hot pressing. While the skutterudite phase was successfully synthesized, a small amount of the secondary phase (FeSb2) was observed. According to the scanning electron microscope analysis, (Ce,Pr)Sb-2 phases were also observed for Co-substituted specimens (x = 0.5). The electrical conductivity decreased with increasing temperature, implying a degenerate semiconductor behavior, and also decreased with increasing Co contents. All specimens showed p-type characteristics having positive signs of the Hall coefficient and the Seebeck coefficient. The Seebeck coefficient increased with increasing temperature and reached a maximum value at 823 K. The power factor (PF) increased with decreasing Co content and Ce0.75Pr0.25 Fe4Sb12 showed a peak value of PF = 3.2 mW m(-1) K-2 at 823 K. The electronic thermal conductivity decreased with increasing Co contents and the lattice thermal conductivity decreased with decreasing Ce and Co contents at high temperature. The thermal conductivity increased at temperatures above 623 K due to bipolar conduction. The dimensionless figurea of pound merit (ZT) showed a maximum value of ZT = 0.84 at 823 K for Ce0.25Pr0.75Fe4Sb12.</P>

Pinoresinol-4,4'-di-O-β-d-glucoside from Valeriana officinalis root stimulates calcium mobilization and chemotactic migration of mouse embryo fibroblasts

Do, K.H.,Choi, Y.W.,Kim, E.K.,Yun, S.J.,Kim, M.S.,Lee, S.Y.,Ha, J.M.,Kim, J.H.,Kim, C.D.,Son, B.G.,Kang, J.S.,Khan, I.A.,Bae, S.S. G. Fischer 2009 Phytomedicine Vol.16 No.6

Lignans are major constituents of plant extracts and have important pharmacological effects on mammalian cells. Here we showed that pinoresinol-4,4'-di-O-β-d-glucoside (PDG) from Valeriana officinalis induced calcium mobilization and cell migration through the activation of lysophosphatidic acid (LPA) receptor subtypes. Stimulation of mouse embryo fibroblast (MEF) cells with 10μM PDG resulted in strong stimulation of MEF cell migration and the EC<SUB>50</SUB> was about 2μM. Pretreatment with pertussis toxin (PTX), an inhibitor of G<SUB>i</SUB> protein, completely blocked PDG-induced cell migration demonstrating that PDG evokes MEF cell migration through the activation of the G<SUB>i</SUB>-coupled receptor. Furthermore, pretreatment of MEF cells with Ki16425 (10μM), which is a selective antagonist for LPA<SUB>1</SUB> and LPA<SUB>3</SUB> receptors, completely blocked PDG-induced cell migration. Likewise, PDG strongly induced calcium mobilization, which was also blocked by Ki16425 in a dose-dependent manner. Prior occupation of the LPA receptor with LPA itself completely blocked PDG-induced calcium mobilization. Finally, PDG-induced MEF cell migration was attenuated by pretreatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor such as LY294002. Cells lacking downstream mediator of PI3K such as Akt1 and Akt2 (DKO cells) showed loss of PDG-induced migration. Re-expression of Akt1 (but not Akt2) completely restored PDG-induced DKO cell migration. Given these results, we conclude that PDG is a strong inducer of cell migration. We suggest that the pharmacological action of PDG may occur through the activation of an LPA receptor whereby activation of PI3K/Akt signaling pathway mediates PDG-induced MEF cell migration.

MOA-2010-BLG-073L: AN M-DWARF WITH A SUBSTELLAR COMPANION AT THE PLANET/BROWN DWARF BOUNDARY

Street, R. A.,Choi, J.-Y.,Tsapras, Y.,Han, C.,Furusawa, K.,Hundertmark, M.,Gould, A.,Sumi, T.,Bond, I. A.,Wouters, D.,Zellem, R.,Udalski, A.,Snodgrass, C.,Horne, K.,Dominik, M.,Browne, P.,Kains, N.,Br IOP Publishing 2013 The Astrophysical journal Vol.763 No.1

<P>We present an analysis of the anomalous microlensing event, MOA-2010-BLG-073, announced by the Microlensing Observations in Astrophysics survey on 2010 March 18. This event was remarkable because the source was previously known to be photometrically variable. Analyzing the pre-event source light curve, we demonstrate that it is an irregular variable over timescales >200 days. Its dereddened color, (V - I)(S),(0), is 1.221 +/- 0.051 mag, and from our lens model we derive a source radius of 14.7 +/- 1.3 R-circle dot, suggesting that it is a red giant star. We initially explored a number of purely microlensing models for the event but found a residual gradient in the data taken prior to and after the event. This is likely to be due to the variability of the source rather than part of the lensing event, so we incorporated a slope parameter in our model in order to derive the true parameters of the lensing system. We find that the lensing system has a mass ratio of q = 0.0654 +/- 0.0006. The Einstein crossing time of the event, t(E) = 44.3 +/- 0.1 days, was sufficiently long that the light curve exhibited parallax effects. In addition, the source trajectory relative to the large caustic structure allowed the orbital motion of the lens system to be detected. Combining the parallax with the Einstein radius, we were able to derive the distance to the lens, D-L = 2.8 +/- 0.4 kpc, and the masses of the lensing objects. The primary of the lens is an M-dwarf with M-L,M-1 = 0.16 +/- 0.03 M-circle dot, while the companion has M-L,M-2 = 11.0 +/- 2.0 M-J, putting it in the boundary zone between planets and brown dwarfs.</P>

Supplementation of oil-based inactivated H9N2 vaccine with M2e antigen enhances resistance against heterologous H9N2 avian influenza virus infection

Park, J.K.,Lee, D.H.,Cho, C.H.,Yuk, S.S.,To, E.O.,Kwon, J.H.,Noh, J.Y.,Kim, B.Y.,Choi, S.W.,Shim, B.S.,Song, M.K.,Lee, J.B.,Park, S.Y.,Choi, I.S.,Song, C.S. Elsevier Scientific Pub. Co 2014 Veterinary microbiology Vol.169 No.3

Avian influenza virus (AIV) subtype H9N2 has been evolving rapidly and vaccine escape variants have been reported to cause circulation of infections and economic losses. In the present study, we developed and evaluated ectodomain of the AIV matrix 2 (M2e) protein as a supplementing antigen for oil-based inactivated H9N2 vaccine to increase resistance against vaccine escape variants. AIV H9N2 M2e antigen was expressed in Escherichia coli and supplemented to inactivated H9N2 oil emulsion vaccine. Specific pathogen-free chickens received a single injection of inactivated H9N2 oil emulsion vaccines with or without M2e supplementation. At three weeks post vaccination, hemagglutination inhibition tests and enzyme-linked immunosorbent assays were performed to determine serological immune responses. Challenge study using a vaccine escape H9N2 variant was performed to evaluate the efficacy of M2e supplementation. M2e antigen supplemented in oil emulsion vaccine was highly immunogenic, and a single M2e-supplemented vaccination reduced challenge virus replication and shedding more effectively than non-supplemented vaccination.

Effect of Antimuscarinic Autoantibodies in Primary Sjögren’s Syndrome

Kim, N.,Shin, Y.,Choi, S.,Namkoong, E.,Kim, M.,Lee, J.,Song, Y.,Park, K. SAGE Publications 2015 Journal of dental research Vol.94 No.5

<P>The presence of functional autoantibodies against the muscarinic type 3 receptor (M3R) has been reported in primary Sjogren's syndrome (pSS). However, the pathogenic role of these autoantibodies in pSS development remains to be elucidated. In this experiment, we investigated a pathologic role of pSS autoantibodies (pSS IgG) associated with downregulation of the major histocompatibility complex I (MHC I) molecule with M3R through internalization. Anti-M3R autoantibodies in purified control and pSS IgG were detected using 4 synthesized cyclic M3R peptides by enzyme-linked immunosorbent assay. The binding reactivity of pSS IgG to M3R in situ was analyzed by a dual immunostaining method. Surface expression, interaction, and internalization of M3R with MHC I were analyzed by immunofluorescence confocal microscopy and biochemical assays. Synthetic cyclic peptides M3RP(205-221) and M3RP(520-527) showed significantly high reactivity with pSS IgG compared to the control IgG or the other 3 peptides (P < 0.05). Significantly high reactivity of pSS IgG to M3R in situ was observed. PSS IgG increased the interaction of membrane M3R with MHC I and induced their internalization in primary human submandibular gland cells. The pSS IgG-induced internalization of M3R with MHC I was significantly inhibited by the cholesterol-sequestering drug filipin. Our novel findingnamely, strong downregulation of the membrane MHC I with M3R through internalization of the cholesterol-rich microdomain associating with anti-M3R autoantibodiescould be an important mechanism contributing to the impaired salivation seen in pSS and linking secretory hypofunction to autoimmune pathogenesis.</P>

가토에서 CCNU 로 유발된 골수 저형성증의 혈액학적 소견과 골수스캔의 변화에 관한 연구

김승택,이명철,최두혁,고창순,김병국,이문호,박선양,최성재,김노경,최영희 대한내과학회 1986 대한내과학회지 Vol.30 No.1

To understand systematically the hematological changes including the bone marrow changes in chemotherapeutic agent-induced bone marrow hypoplasia and to define the relationship between hematological and bone marrow scan findings and prognosis of the hypoplasia, CCNU (lomustine) was given orally to 44 rabbits to induce hypoplasia of the bone marrow. And serial changes of peripheral blood and bone marrow findings and (111)In Cl(3)((111)In scan)/(99m)Tc tin colloid bone marrow scan((99m)Tc scan) were checked before and after induction of hypoplasia. With assessment of 28 evaluable rabbits, the following results were obtained: 1) Significant hypoplasia of the bone marrow developed around day 4 of CCNU administration and recovered around day 14(cellularity 51,4±13.5% and 24. 5±14.97o before and after CCNU respectively, p<005). Megakaryocyte count was significantly depressed from 95.67,26% to 36.7$gt;31.82%(P$lt;0.005). M: E ratio was decreased from 162±1.19 to 0, 0.5$lt;0. 43(p±0.005). Shift to left(475), maturation a(40%), naked nucleus and degenerated cells(20%), increase of lymphocytes(47%), monocytes and reticulum cells were also found. 2) The uptake ratio of the 99(m)Tc tin colloid bone marrow scan was markedly increased in contrast to the depression of the bone marrow(4.4±2.12 and 14.1±7.06 before and 4 days after CCNU, respectively, p40. 005). Tc scan uptake ratio was inversely related to the cellularity(r=-0.442, p$lt;0.05) and megakaryocyte number of the bone marrow(r= 0.89, p< 0.01) and peripheral blood granulocyte (r = 0. 54. Pg0.01) and platelet count(r=0.40, p$lt;0.05). There was not significant correlation between (111)In scan uptake ratio and hematologic parameters. 3) The amplitude of the change of the (99m)Tc scan uptake ratio was significantly related to the prognosis of the rabbits with experimentally induced hypoplasia of the bone marrow(dead 5.1±2,67, survivors 2,5±0.96, P<0.01). The change of the (111)In scan uptake ratio was not related to the prognosis of these rabbits. In experimentally induced rrow hypoplasia, morphologic changes in addition to the numerical changes of the bone marrow elements were observed. And serial (99)Tc scan of the bone marrow appears to be helpful assessing the severity and predicting the outcome of bone marrow hypoplasia.

Metabolic characterization of (1-(5-fluoropentyl)-1H-indol-3-yl)(4-methyl-1-naphthalenyl)-methanone (MAM-2201) using human liver microsomes and cDNA-overexpressed cytochrome P450 enzymes

Kong, T. Y.,Kim, J. H.,Choi, W. G.,Lee, J. Y.,Kim, H. S.,Kim, J. Y.,In, M. K.,Lee, H. S. Springer Science + Business Media 2017 Analytical and bioanalytical chemistry Vol.409 No.6

<P>MAM-2201 is a synthetic cannabinoid that is increasingly found in recreational drug abusers and cases of severe intoxication. Thus, characterization of the metabolic pathways of MAM-2201 is necessary to predict individual pharmacokinetics and toxicity differences, and to avoid toxic drug-drug interactions. Collectively, 19 phase 1 metabolites of MAM-2201 were identified using liquid chromatography-Orbitrap mass spectrometry following human liver microsomal incubations in the presence of NADPH: 7 hydroxy-MAM-2201 (M1-M7), 4 dihydroxy-MAM-2201 (M8-M11), dihydrodiol-MAM-2201 (M12), N-(5-hydroxypentyl)-MAM-2201 (M13), hydroxy-M13 (M14), N-dealkyl-MAM-2201 (M15), 2 hydroxy-M15 (M16, M17), MAM-2201 N-pentanoic acid (M18), and hydroxy-M18 (M19). On the basis of intrinsic clearance values in human liver microsomes, hydroxy-MAM-2201 (M1), N-(5-hydroxypentyl)-MAM-2201 (M13), and hydroxy-M13 (M14) were the major metabolites. Based on an enzyme kinetics study using human cDNA-expressed cytochrome P450 (CYP) enzymes and an immunoinhibition study using selective CYP antibodies in human liver microsomes, CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 enzymes were responsible for MAM-2201 metabolism. The CYP3A4 enzyme played a prominent role in MAM-2201 metabolism, and CYP1A2, CYP2B6, CYP2C8, and CYP2C9 enzymes played major roles in the formation of some metabolites. MAM-2201 is extensively metabolized by multiple CYP enzymes, indicating that MAM-2201 and its metabolites should be used as markers of MAM-2201 abuse and toxicity.</P>

Multistage symmetry breaking in the breathing pyrochlore latticeLi(Ga,In)Cr4O8

Lee, S.,Do, S.-H.,Lee, W.-J.,Choi, Y. S.,Lee, M.,Choi, E. S.,Reyes, A. P.,Kuhns, P. L.,Ozarowski, A.,Choi, K.-Y. American Physical Society 2016 Physical Review B Vol.93 No.17

<P>We present magnetic susceptibility, dielectric constant, high-frequency electron spin resonance, Li-7 nuclear magnetic resonance, and zero-field muon spin relaxation measurements of LiACr(4)O(8) (A = Ga, In), towards realizing a breathing pyrochlore lattice. Unlike the uniform pyrochlore ZnCr2O4 lattice, both the In and the Ga compounds feature two-stage symmetry breaking: a magnetostructural phase transition with subsequent antiferromagnetic ordering. We find a disparate symmetry breaking process between the In and the Ga compounds, having different degrees of bond alternation. Our data reveal that the Ga compound with moderate bond alternation shows the concomitant structural and magnetic transition at T-S = 15.2 K, followed by the magnetic ordering at T-m = 12.9 K. In contrast, the In compound with strong bond alternation undergoes a thermal crossover at T* approximate to 20.1 K from a tetramer singlet to a dimer singlet or a correlated paramagnet with a separate weak magnetostructural transition at T-S = 17.6 K and the second antiferromagnetic ordering at T-m = 13.7 K. This suggests that the magnetic phases and correlations of the breathing pyrochlore lattice can be determined from the competition between bond alternation and spin-lattice coupling, thus stabilizing long-range magnetic ordering against a nonmagnetic singlet.</P>