Activation of succinate dehydrogenase by 2,4-dinitrophenol-a competitive inhibitor

1.The reported inhibition of the succinate oxidase system at high concentrations of dinitrophenol, considered to be at the primary dehydrogenase level, is now confirmed by measuring the activity of succinate dehydrogenase (succinate:(acceptor) oxidoreductase, EC 1.3.99.1) in the presence of dinitrophenol, using the dye reduction method. 2. 2. The results indicate that the inhibition of substrate-activated succinate dehydrogenase by dinitrophenol is competitive. 3. 3. Low concentrations of dinitrophenol inhibited the basal activity, while at higher concentrations the kinetics were complicated by an apparent activation. 4. 4. Preincubation of mitochondria with dinitrophenol stimulated the enzyme activity, a phenomenon shown by succinate and competitive inhibitors. This activation was very rapid at 37°, compared to that by succinate; activation by dinitrophenol was observed even at 25°, under conditions where succinate had no effect. 5. 5. Repeated washing of the activated mitochondrial samples with the sucrose homogenizing medium reduced the succinate-stimulated activity to the basal level, but only partially reversed the dinitrophenol activation. 6. 6. The relevance of this activation phenomenon to the physiological modulation of this enzyme system is discussed.

Cloning, expression, purification and preliminary X-ray crystallographic studies of 2-methylisocitrate lyase from Salmonella typhimurium. Acta Crystallogr

In Salmonella typhimurium, propionate is oxidized to pyruvate via the 2-methylcitric acid cycle. The last step of this cycle, the cleavage of 2-methylisocitrate to succinate and pyruvate, is catalysed by 2-methylisocitrate lyase (EC 4.1.3.30). Methylisocitrate lyase (molecular weight 32 kDa) with a C-terminal polyhistidine affinity tag has been cloned and overexpressed in Escherichia coli and purified and crystallized under different conditions using the hanging-drop vapour-diffusion technique. Crystals belong to the orthogonal space group P2(1)2(1)2(1), with unit-cell parameters a = 63.600, b = 100.670, c = 204.745 Angstrom. A complete data set to 2.5 Angstrom resolution has been collected using an image-plate detector system mounted on a rotating-anode X-ray generator.

Identification of the active-site peptide of 2,3-dihydroxybenzoic acid decarboxylase from Aspergillus oryzae

The non-oxidative decarboxylation of aromatic acids is a poorly understood reaction. The transformation of 2,3-dihydroxybenzoic acid to catechol in the fungal metabolism of indole is a prototype of such a reaction. 2,3-Dihydroxybenzoic acid decarboxylase (EC 4.1.1.46) which catalyzes this reaction was purified to homogeneity from anthranilate induced cultures of Aspergillus oryzae using affinity chromatography. The enzyme did not require cofactors like NAD(+), PLP, TPP or metal ions for its activity. There was no spectral evidence for the presence of enzyme bound cofactors. The preparation, which was adjudged homogeneous by the criteria of SDS-PAGE, sedimentation analysis and N-terminal analysis, was characterized for its physicochemical and kinetic parameters. The enzyme was inactivated by group-specific modifiers like diethyl pyrocarbonate (DEPC) and N-ethylmaleimide (NEM). The kinetics of inactivation by DEPC suggested the presence of a single class of essential histidine residues, the second order rate constant of inactivation for which was 12.5 M(-1) min(-1). A single class of cysteine residues was modified by NEM with a second order rate constant of 33 M(-1) min(-1). Substrate analogues protected the enzyme against inactivation by both DEPC and NEM, suggesting the Location of the essential histidine and cysteine to be at the active site of the enzyme. The incorporation of radiolabelled NEM in a differential labelling experiment was 0.73 mol per mol subunit confirming the presence of a single essential cysteine per active-site. Differentially labelled enzyme was enzymatically cleaved and the peptide bearing the label was purified and sequenced. The active-site peptide LLGLAETCK and the N-terminal sequence MLGKIALEEAFALPRFEEKT did not bear any similarity to sequences reported in the Swiss-Prot Protein Sequence Databank, a reflection probably of the unique primary structure of this novel enzyme. The sequences reported in this study will appear in the Swiss-Prot Protein Sequence Databank under the accession number P80402.

Unimolecular HCl elimination from 1,2-dichloroethane: A single pulse shock tube and ab initio study

Thermal decomposition of 1,2-dichloroethane (1,2-DCE) has been studied in the temperature range of 10501175 K behind reflected shock waves in a single pulse shock tube. The unimolecular elimination of HCl is found to be the major channel through which 1,2-DCE decomposes under these conditions. The rate constant for the unimolecular elimination of HCl from 1,2-dichloroethane is found to be 10(13.98+/-0.80) exp(-57.8+/-2.0/RT) s(-1), where the activation energy is given in kcal mol(-1) and is very close to that value for CH3CH2Cl (EC). Ab initio (HF and MP2) and DFT calculations have been carried out to find the activation barrier and the structure of the transition state for this reaction channel from both EC and 1,2-DCE. The preexponential factors calculated at various levels of theory (BF/6-311++G**, MP2/6-311++G**, and B3LYP/6-311++G**) are (approximate to10(15) s(-1)) significantly larger than the experimental results. If the torsional mode in the ground state is treated as free internal rotation the preexponential factors reduce significantly, giving excellent agreement with experimental values. The DFT results are in excellent (fortuitous?) agreement with the experimental value for activation energy for 1,2-DCE while the MP2 and HF results seem to overestimate the barrier. However, DFT results for EC is 4.5 kcal mol(-1) less than the previously reported experimental values. At all levels, theory predicts an increase in HCI elimination barrier on beta-Cl substitution on EC.

Aza-induced cardiomyocyte differentiation of P19 EC-cells by epigenetic co-regulation and ERK signaling

Stem cells in cell based therapy for cardiac injury is being potentially considered. However, genetic regulatory networks involved in cardiac differentiation are not clearly understood. Among stem cell differentiation models, mouse P19 embryonic carcinoma (EC) cells, are employed for studying (epi)genetic regulation of cardiomyocyte differentiation. Here, we comprehensively assessed cardiogenic differentiation potential of 5-azacytidine (Aza) on P19 EC-cells, associated gene expression profiles and the changes in DNA methylation, histone acetylation and activated-ERK signaling status during differentiation. Initial exposure of Aza to cultured EC-cells leads to an efficient (55%) differentiation to cardiomyocyte-rich embryoid bodies with a threefold (16.8%) increase in the cTnI(+) cardiomyocytes. Expression levels of cardiac-specific gene markers i.e., Isl-1, BMP-2, GATA-4, and alpha-MHC were up-regulated following Aza induction, accompanied by differential changes in their methylation status particularly that of BMP-2 and alpha-MHC. Additionally, increases in the levels of acetylated-H3 and pERK were observed during Aza-induced cardiac differentiation. These studies demonstrate that Aza is a potent cardiac inducer when treated during the initial phase of differentiation of mouse P19 EC-cells and its effect is brought about epigenetically and co-ordinatedly by hypo-methylation and histone acetylation-mediated hyper-expression of cardiogenesis-associated genes and involving activation of ERK signaling.

Evaluación de dosis del herbicida Pyribenxozim (Pyanchor 5 ec) aplicados en post emergencia temprana, para el control de arvenses en época seca en el cultivo de arroz (Oryza sativa L) Timal, Tipitapa, 2006

El presente estudio se realizó en febr ero del año 2006, en Tipi tapa, municipio del departamento de Managua, Km 26 carretera norte. En la zona arrocera el Timal, finca del pivote 34, del señor Freddy González, con el objetivo de ev aluar la eficiencia de tres dosis del herbicida Pyribenxozim (pyanc hor 5 EC) aplicado en post emergencia temprana para el control de arvenses en el cultivo de arroz (Oryza sativa L), de riego en época seca. El experimento se estableció en un lote comercial, en un Diseño de Bloque Completo al Azar (BCA), con cinco tratamientos y cuatro repeticiones. Las variables evaluadas en este estudio fueron: composición florística, cobertura, Efectividad del herbicida sobre las arvenses selectividad (Fitotoxicidad) y rendimiento. Las dosis del producto evaluado en el ensayo fueron: T1- (Pyribenxozim 5 EC 0.8 l/ha), T2 - (Pyribenxozim 5 EC 1.0 l/ha), T3 - (Pyribenxozim 5 EC 1.2 l/ha), T4 - (Testigo comercial Bispiribac Sodico (Nominee) 40 SC 1.0 l/ha) (muy utilizado en la zona) y T5 - (Testigo absoluto). En los resultados, se identificaron las arvenses que predominaban en el cultivo, las principales familias predominantes fueron las Poaceae. Los mayores porcentajes de c obertura se encontraron en T1- (Pyribenxozim 5 EC 0.8 l/ha) y el T5 - (Testigo absoluto). Los tres tratamientos evaluados de pyanchor 5 EC controlaron eficientemente las principales arvenses presentes. Sin embargo el mejor resultado sobre el detrimento de las arvenses fue la dosis de 1.2 l/ ha de Pyanchor 5 EC en la que se encontró un 97 % de control. En cuanto a la efectividad sobre el tipo de especies, se observó, que ninguna de las dosis estudiadas logro afectar a las especies Leptochloa filiformis L y Eclipta alba L mismo efecto se observó en el tratamiento en el que se uso el producto comercial Bispir ibac Sodico (Nominee) 40 SC 1.0 l/ha. El herbicida Pyribenxozim (pyanchor 5 EC), es un producto selectivo con respecto al cultivo de arroz. Según esto s resultados, en el rendimien to del cultivo ninguno de los tratamientos afecto su calidad y cantidad según características de la variedad. En cuanto a la probación y registro del producto por el MAG FOR, basado en los resultados obtenidos este actualmente fue aceptado y esta siendo aplicado por los productores de arroz.

30. Sitzung des Codex Alimentarius Komitees für Fische und Fischerzeugnisse in Agadir, Marokko, vom 28.9. bis 2.10.2009

The Codex Committee on Fish and Fishery Products held its 30th Session in Agadir, Morocco from 28 September to 2 October 2009, at the kind invitation of the Government of Morocco. The Session was chaired by Dr Bjørn Røthe Knudsen, Regional Director of the Norwegian Food Safety Authority. The Session was attended by 218 delegates representing 78 Member States, one Member Organization (EC) and 1 international organization.

Effect of diazinon 60 EC on Anabas testudineus, Channa punctatus and Barbodes gonionotus

Anabas testudineus, Channa punctatus and Barbodes gonionotus were exposed to 5.62, 6.25, 6.87, 7.50, 8.12 and 8.75 ppm; 1.13, 2.26, 3.39, 4.52, 5.65 and 6.78 ppm; and 2.00, 2.50, 3.00, 3.50, 4.00 and 4.50 ppm of Diazinon 60 EC, respectively. The median lethal concentration (LC50) values of Diazinon 60 EC on A. testudineus, C. punctatus and B. gonionotus were 6.55, 3.09 and 2.72 ppm for 96 hrs of exposure. The fish species showed several abnormal behaviors which included restlessness, arena movements, loss of equilibrium, increased opercular activities, strong spasm, paralysis and sudden quick movements during the exposure. For histopathological studies, A. testudineus, C. punctatus and B. gonionotus were exposed for 7 days to sublethal concentrations of 1.13 and 3.75 ppm; 1.13 and 2.26 ppm; and 1.13 and 2.26 ppm of Diazinon 60 EC, respectively. Hypertrophy, necrosis and pyknosis of hepatocytes, pyknosis and degenerative changes such as necrosis of tubular and haematopoietic cells of kidney were the major histopathological effects.

~(133)Sm的(EC+β~+)衰变和~(133)Pr的1.1s同质异能态

通过96 Ru( 4 0 Ca ,ln2p)反应 ,采用氦喷嘴带传输系统和X γ与γ γ符合测量方法 ,首次建议了133Sm的简单的 (EC + β+)衰变纲图 .由于Ru靶中含有98— 10 2 Ru的成分 ,同时产生了133Pr,并首次测定了133Pr的 1 1 / 2 - 同质异能态的寿命为( 1 .1± 0 .2 )s.用单粒子模型提取了131,133,135 ,137Pr的 1 1 / 2 - 同质异能态的约化跃迁几率的实验值 ,并与Weisscopf近似估计进行了比较 .

~(130)Ce(EC+β~+)衰变研究及其放射化学分离

利用兰州SFC加速的16O束轰击同位素118Sn ,由熔合蒸发 4n反应产生目标核13 0 Ce。为了消除本底干扰并指定13 0 Ce核 ,采用溶剂萃取法对He - jet带传输系统从靶室传输出来的反应产物进行了离线分离与纯化。将目标核13 0 Ce从大量的靶材料、反应产物及母核中分离出来 ,快速制成薄源后在铅室中进行γ单谱测量及X -γ、γ -γ符合测量。从化学分离后的产物中观察到了半衰期为 2 2 .9min的 10 8条γ射线 ,其中 10 7条是新发现的 ,该活性被指定为13 0 Ce。在此基础上 ,进一步研究这些γ线的级联关系 ,建立了缺中子同位素13 0 Ce较完整的 (EC + β+ )衰变纲图。为118Sn(16O ,4n) 13 0 Ce反应体系建立的放化分离流程的分离时间仅 10min ,化学产额大于70 %。化学分离除去 98%以上的核反应生成的13 0 La ,对其它杂质的去污完全满足13 0 Ce(EC + β+ )衰变研究的要求。

缺中子核素~(129)Ce的(EC/β~+)衰变

利用 10 2MeV16O6+束轰击同位素靶117Sn ,通过熔合蒸发 4n反应产生核素12 9Ce.由氦喷嘴快速带传输系统将反应产物送到低本底区 .通过化学分离来制备待测的Ce样品 ,与此同时用16O束轰击117Sn的两种相邻的同位素靶118Sn和116Sn ,并比较上述 3种反应中的产物来进一步区分元素Ce的不同的同位素 .结果一种半衰期为 4.1min的活性被鉴定为12 9Ce.基于X -γ -t和γ -γ -t符合测量 ,建议了包括 5 1条射线在内的12 9Ce的 (EC/ β+)衰变纲图 .其中 ,12 9Ce基态直接馈送到12 9La基态的份额 ( 2 6± 7) %是用观测到的12 9La衰变的 2 78.6keV的γ射线的生长和衰变曲线估计出来的 .另外还给出了用La Kα X射线和6 8.2keVγ射线开门的γ谱以及典型的衰变γ射线的时间谱

EC/beta(+) decay of six medium-heavy nuclei

Previous experimental results of (EC+beta(+)) decay for the medium-heavy nuclei reported by our group since 1996, including Er-153, Yb-157, Fr-209, Ce-128, Ce-130, and Pr-128 have been briefly summarized. The observed low-lying states in their daughter nuclei have been reviewed in a systematic way and compared with different model calculations. Finally, some questions have been put forward for further study and discussion.