Interaction between tryptophan‐Sm(III) complex and DNA with the use of a acridine orange dye fluorophor probe

The interaction of the Trp–Sm(III) complex with herring sperm DNA (hs‐DNA) was investigated with the use of acridine orange (AO) dye as a spectral probe for UV‐vis spectrophotometry and fluorescence spectroscopy. The results showed that the both the Trp–Sm(III) complex and the AO molecule could intercalate into the double helix of the DNA. The Sm(III)–(Trp)₃ complex was stabilized by intercalation into the DNA with binding constants: K^?_25°C = 7.14 × 10⁵ L·mol^?1 and K^?_37°C = 5.28 × 10⁴ L·mol^?1, and it could displace the AO dye from the AO–DNA complex in a competitive reaction. Computation of the thermodynamic functions demonstrates that Δ_rH_m^? is the primary driving power of the interaction between the Sm(III)(Trp)₃ complex and the DNA. The results from Scatchard and viscometry methods suggested that the interaction mode between the Sm(III)(Trp)₃ complex and the hs‐DNA is groove binding and weak intercalation binding. Copyright © 2015 John Wiley & Sons, Ltd.     

The reaction between cytochrome C1 and cytochrome C

The kinetics of electron transfer between the isolated enzymes of cytochrome c₁ and cytochrome c have been investigated using the stopped-flow technique. The reaction between ferrocytochrome c₁ and ferricytochrome c is fast; the second-order rate constant (k₁) is 3.0 · 10⁷ M^?1 · s^?1 at low ionic strength (I = 223 mM, 10°C). The value of this rate constant decreases to 1.8 · 10⁵ M^?1 · s^?1 upon increasing the ionic strength to 1.13 M. The ionic strength dependence of the electron transfer between cytochrome c₁ and cytochrome c implies the involvement of electrostatic interactions in the reaction between both cytochromes. In addition to a general influence of ionic strength, specific anion effects are found for phosphate, chloride and morpholinosulphonate. These anions appear to inhibit the reaction between cytochrome c₁ and cytochrome c by binding of these anions to the cytochrome c molecule. Such a phenomenon is not observed for cacodylate. At an ionic strength of 1.02 M, the second-order rate constants for the reaction between ferrocytochrome c₁ and ferricytochrome c and the reverse reaction are k₁ = 2.4 · 10⁵ M^?1 · s^?1 and k_?1 = 3.3 · 10⁵ M^?1 · s^?1, respectively (450 mM potassium phosphate, pH 7.0, 1% Tween 20, 10°C). The ‘equilibrium’ constant calculated from the rate constants (0.73) is equal to the constant determined from equilibrium studies. Moreover, it is shown that at this ionic strength, the concentrations of intermediary complexes are very low and that the value of the equilibrium constant is independent of ionic strength. These data can be fitted into the following simple reaction scheme: cytochrome c²⁺₁ + cytochrome c³⁺ai cytochrome c³⁺₁ + cytochrome c²⁺.     

THF INFLUENCE OF IRRADIANCE ON THF BIOCHEMICAL COMPOSITION OF THE PRYMNESIOPHYTE ISOCHRYSIS SP. (CLONE T-ISO)1

The effects of irradiance on the biochemical composition of the prymnesiophyte microalga, Isochrysis sp. (Parke; clone T-ISO), a popular species for mariculture, were examined. Cultures were grown under a 12:12 h light: dark (L:D) regime at five irradiances ranging from 50 to 1000 μE·m ²·s^?1 and harvested at late-logarithmic phase for analysis of biochemical composition. Gross composition varied aver the range of irradiances. The highest levels of protein were present in cells from cultures grown at 100 and 250 μE·m ³·s¹, and minimum levels of carbohydrate and lipid occurred at 50 μE·m^?2·s^?1. Because the cell dry weight was reduced at lower irradiances, different trends were evident when results were expressed as percentage of dry weights. Protein percentages were highest at Wand 100 μE·m^?2·s^?1 and carbohydrate at 100 μE·m^?2·s^?1. The composition of amino acids did not differ over the range of irradiances. Glutamate and aspartate were always present in high proportions (9.0–13.5%); histidine. methionine, tryptophan, cystine, and hydroxy-proline were minor constituents (0.0–2.6%). Glucose was the predominant sugar in all cultures, ranging from 23.0% (50 μE·m^?2·s^?1) to 45.0% (100 μE·m^?2·s^?1) of total polysaccharide. No correlation was found between the proportion of any of the sugars and irradiance. The proportions of the lipid class components and fatty acids showed little change with irradiance. The main fatty acids were 14:0, 16:0, 16:1(n-7), 18:1(n-9), 18:3(n-3). 18:4(n-3), 18:5(n-3), and 22:6(n-3). Proportions of 22: 6(n-3) increased, whereas l8:3(n-3). 18:3(n-6). and 18:4(n-3) decreased, with increasing irradiance. Pigment concentrations were highest in cultures grown at 50 μE·m^?2·s^?1, except for fucoxanthin and diadinoxanthin (100 μE·m^?2·s^?1). The concentrations of accessory pigments correlated with chlorophyll a, which decreased in concentration with increasing irradiance. On the basts of biochemical composition, an irradiance of 100 μE·m^?1·s^?1 (12:12 h L:D cycle)for the culture of Isochrysis sp. (clone T-ISO) may provide optimal nutritional value for maricultured animals, although feeding trials are now necessary to substantiate this.     

PHYSIOLOGICAL RESPONSES OF ANABAENA VARIABILIS (CYANOPHYCEAE) TO INSTANTANEOUS EXPOSURE TO VARIOUS COMBINATIONS OF LIGHT INTENSITY AND TEMPERATURE1

Light intensity and temperature interactions have a complex effect on the physiological process rates of the filamentous bluegreen alga Anabaena variabilis Kütz. The optimum temperature for photosynthesis increased with increasing light intensity from 10°C at 42 μE·m^?2·s^?1 to 35°C at 562 μE·m^?2·s^?1. The light saturation parameter, I_K, increased with increasing temperatures. The maximum photosynthetic rate (2.0 g C·g dry wt.^?1·d^?1) occurred at 35°C and 564 μE·m^?2·s^?1. At 15°C, the maximum rate was 1.25 g C·g dry wt.^?1·d^?1 at 332 μE·m^?2·s^?1. The dark respiration rate increased exponentially with temperature. Under favorable conditions of light intensity and temperature the percent of extracellular release of dissolved organic carbon was less than 5% of the total C fixed. This release increased to nearly 40% under combinations of low light intensity and high temperature. A mathematical model was developed to simulate the interaction of light intensity and temperature on photosynthetic rate. The interactive effects were represented by making the light-saturation parameters a function of temperature.     

The binding of arsenazo III to cell components

The Ca²⁺ indicator, arsenazo III, binds to subcellular fractions of rabbit skeletal muscle with sufficient affinity that in living muscle containing 1–2 mM arsenazo III, the estimated free arsenazo III concentration is only 50–200 μM; 80–90% of the bound arsenazo III is associated with soluble proteins.The binding of arsenazo III to soluble proteins decreases the optical response of the dye to Ca²⁺; this is due to a decrease in the affinity of the protein-bound dye for Ca²⁺. Approximately half of the bound arsenazo III is released from the particulate fraction and soluble proteins upon addition of 5 mM Ca²⁺, suggesting that the Ca-arsenazo complex has lower affinity for the protein binding sites than the free dye.The Ca²⁺ binding to the soluble protein fraction of rabbit skeletal muscle is attributable largely to its parvalbumin content.     

Stacking interactions of ethidium bromide bound to a polyphosphate and phage DNA in situ

Ethidium bromide forms spectroscopically detectable aggregates in aqueous solution and at a high dye concentration larger than 1 × 10^?3 moles/ρ. At moderate concentration in the order of 1 × 10^?4 moles/ρ the dye interacts with inorganic polyphosphate Graham salt and with phage sd DNA in situ by formation of stacking complexes. Maximal stacking was found at a phosphate to dye ratio, P/D, of approximately 1 for Graham salt and 1.5–2 for phages. In going to a higher P/D ratio Graham salt dye complex dissociates again and free dye reappears, while phage dye binding changes from stacking (type II complex) to intercalation (type I complex). Stacking is accompanied by a decrease and intercalation by an increase of relative fluorescence intensity with respect to free dye. However, both binding types lead to hypechromism and a red shift of the dye absorption band in the visible spectral region. Thus spectral behavior of ethidium aggregates deviate clearly from that known for other dyes, i.e., acridines.     

STUDY ON BIOLOGICAL CHARACTERISTICS OF HETEROTROPHIC MARINE MICROALGA—SCHIZOCHYTRIUM MANGROVEI PQ6 ISOLATED FROM PHU QUOC ISLAND,KIEN GIANG PROVINCE,VIETNAM1

Schizochytrium sp. PQ6, a heterotrophic microalga isolated from Phu Quoc (PQ) Island in the Kien Giang province of Vietnam, contains a high amount of docosahexaenoic acid (DHA, C22:6n‐3). In this study, the culture conditions are developed to maximize biomass and DHA production. Nucleotide sequence analysis of partial 18S rRNA gene from genomic DNA showed that PQ6 has a phylogenetic relationship close to Schizochytrium mangrovei Raghu‐Kumar. The highest growth rate and DHA accumulation of this strain were obtained in 6.0% glucose, 1.0% yeast extract, 50% artificial seawater (ASW), and pH 7 at 28°C. In addition, carbon and nitrogen sources could be replaced by glycerol, ammonium acetate, sodium nitrate, or fertilizer N–P–K. Total lipid content reached 38.67% of dry cell weight (DCW), in which DHA and eicosapentaenoic acid (EPA, C20:5n‐3) contents accounted for 43.58% and 0.75% of the total fatty acid (TFA), respectively. In 5 and 10 L fermenters, the cell density, DCW, total lipid content, and maximum DHA yield were 46.50 × 10⁶ cells · mL^?1, 23.7 g · L^?1, 38.56% of DCW, and 8.71 g · L^?1 (in 5 L fermenter), respectively, and 49.71 × 10⁶ cells · mL^?1, 25.34 g · L^?1, 46.23% of DCW, and 11.55 g · L^?1 (in 10 L fermenter), respectively. Biomass of PQ6 strain possessed high contents of Na, I, and Fe (167.185, 278.3, and 43.69 mg · kg^?1 DCW, respectively). These results serve as a foundation for the efficient production of PQ6 biomass that can be used as a food supplement for humans and aquaculture in the future.     

FIRST INSIGHTS INTO IMPROVEMENT OF EICOSAPENTAENOIC ACID CONTENT IN PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE) BY INDUCED MUTAGENESIS1

A strain improvement program was initiated based on mutagenesis with the goal of commercial production of eicosapentaenoic acid (EPA)from EPA-overproducing microalgal strains. Two rounds of mutation and selection were conducted using Phaeodactylum tricornutum Bohlin UTEX #640 as the parent strain. After the first round of mutagenesis, a putative mutant (provisionally labeled 114) was obtained. The EPA content (% of dry weight) of this mutant strain was 37% higher than that of the wild type. 114 was further mutated and another putative mutant (provisionally called II242) was isolated, the EPA content of which was 44% higher than that of the wild type. When cultured with aeration in 1-L flasks, EPA content of the wild type and putative mutants 114 and II242 was, 17.3 mg · g^?1, 31.5mg · g^?1, and 38.6 mg · g^?1 dry biomass, respectively. EPA productivity was 3.48 mg · L^?1· d^?1 4.01 mg · L^?1· d^?1, and 4.98 mg · L^?1· d^?1 respectively. These figures compare favorably with many other promising EPA-producing microorganisms and suggest that the use of a single methodology such as mutation and selection is a way to improve the polyunsaturated fatty acid content of microalgae and other microorganisms.     

PHYSIOLOGICAL VARIATION AND ELECTROPHORETIC BANDING PATTERNS OF GENETICALLY DIFFERENT SEASONAL POPULATIONS OF SKELETONEMA COSTATUM (BACILLARIOPHYCEAE)1

Clones of Skeletonema costatum (Grev.) Cl. isolated from Narragansett Bay, R.I., during different seasons were grouped according to their electrophoretic banding patterns. The growth rates, pg chlorophyll · cell^?1, carbon uptake · cell^?1· h^?1, and carbon uptake · pg chl^?1· h^?1 were measured at 20°C, in a 14:10 h L:D cycle at 180 μE · m^?2· s^?1. Statistically significant sources of variation were found among groups of clones in growth rate, pg chl · cell^?1, and carbon uptake · pg chl^?1· h^?1. It was concluded that there is a significant relationship between the physiological characteristics of clones isolated from populations in different seasons and patterns of genetic variation inferred from the electrophoretic studies. However, genetic diversity detected by banding patterns tends to underestimate the total genetic diversity in natural populations. The groups of clones most common in summer bloom populations had significantly higher growth rates, lower values of pg chl · cell^?1, and higher rates of carbon uptake · pg chl^?1· h^?1 at 20°C than did the group of clones most common in winter bloom populations. However, differences among groups in these parameters at 20°C alone cannot account for the seasonal cycling of genetically variable populations of Skeletonema in Narragansett Bay. The range of growth rates among clones of this species is 0.1–5.0 divisions · d^?1 under a single set of temperature and light conditions. Chlorophyll concentrations range from 0.2–1.7 pg chl · cell^?1 and carbon uptake · pg chl^?1· h^?1 varies by a factor of 7 among clones. The range of physiological variation in this species means that it is difficult to use laboratory studies of single clones to analyze the responses of natural populations of Skeletonema.     

EFFECTS OF HARVEST STAGE AND LIGHT ON THE BIOCHEMICAL COMPOSITION OF THE DIATOM THALASSIOSIRA PSEUDONANA1

The marine diatom Thalassiosira pseudonana (Hustedt, clone 3H) Hasle and Heimdal was cultured under three different light regimes: 100 μmol photon · m^?2· s^?1 on 12:12 h light : dark (L:D) cycles; 50 μmol photon · m^?2· s^?2 on 24:0 h L:D; and 100 μmol photon · m^?2· s^?1 on 24:0 h L:D. It was harvested during logarithmic and stationary phases for analysis of biochemical composition. Across the different light regimes, protein (as % of organic weight) was highest in cells during logarithmic phase, whereas carbohydrate and lipid were highest during stationary phase. Carbohydrate concentrations were most affected by the different light regimes; cells grown under 12:12 h L:D contained 37–44% of the carbohydrate of cells grown under 24:0 h L:D. Cells in logarithmic phase had high proportions of polar lipids (79 to 89% of total lipid) and low triacylglycerol (≤10% of total lipid). Cells in stationary phase contained less polar lipid (48 to 57% of total lipid) and more triacylglycerol (22 to 45% of total lipid). The fatty acid composition of logarithmic phase cells grown under 24:0 h L:D were similar, but the 100 μmol photon · m^?2· s^?1 (12:12 h L:D) cells at the same stage contained a higher proportion of polyunsaturated fatty acids (PUFAs) and a lower proportion of saturated and monounsaturated fatty acids due to different levels of 16:0, 16:1(n-7), 16:4(n-1), 18:4(n-3), and 20:5(n-3). With the onset of stationary phase, cells grown at 100 μmol photon · m^?2· s^?1 (both 12:12 and 24:0 h L:D) increased in proportions of saturated and monounsaturated fatty adds and decreased in PUFAs. Concentrations (% organic or dry weight) of 14:0, 16:0, 16:1(n-7), 20:5(n-3), and 22:6(n-3) increased in cells of all cultures during stationary phase. The amino acid compositions of cells were similar irrespective of harvest stage and light regime. For mariculture, the recommended light regime for culturing T. pseudonana will depend on the nutritional requirements of the animal to which the alga is fed. For rapidly growing bivalve mollusc larvae, stationary-phase cultures grown under a 24:0 h L:D regime may provide more energy by virtue of their higher percentage of carbohydrate and high proportions and concentrations of energy-rich saturated fatty acids.     

Discrimination of nitrogen isotopes during absorption of ammonium and nitrate at different nitrogen concentrations by rice (Oryza sativa L.) plants

Studies of uptake of ionic sources of N by two hydroponically grown rice (Oryza sativa L.) cultivars (paddy‐field‐adapted Koshihikari and dryland‐adapted Kanto 168) showed that the magnitude of the nitrogen isotope fractionation (?) for uptake of NH₄⁺ depended on the concentrations of NH₄⁺ and cultivar (averaging –6·1‰ for Koshihikari and –12·0‰ for Kanto 168 at concentrations from 40 to 200 mmol m^?3 and, respectively, –13·4 and –28·9‰ for the two cultivars at concentrations from 0·5 to 4 mol m^?3). In contrast, the ? for uptake of NO₃^? in similar experiments was almost insensitive to the N concentration, falling within a much narrower range (+3·2‰ to –0·9‰ for Koshihikari and –0·9‰ to –5·1‰ for Kanto 168 over NO₃^? concentrations from 0·04 to 2 mol m^?3). From longer term experiments in which Norin 8 and its nitrate‐reductase deficient mutant M819 were grown with 2 or 8 mol m^?3 NO₃^? for 30 d, it was concluded that the small concentration‐independent isotopic fractionation during absorption of this ion was not related to nitrate reductase activity.     

ROLE OF GLUTAMATE DEHYDROGENASE AND GLUTAMINE SYNTHETASE IN CHLORELLA VULGARIS DURING ASSIMILATION OF AMMONIUM WHEN JOINTLY IMMOBILIZED WITH THE MICROALGAE‐GROWTH‐PROMOTING BACTERIUM AZOSPIRILLUM BRASILENSE1

Enzymatic activities of glutamate dehydrogenase (GDH) and glutamine synthetase (GS) participating in the nitrogen metabolism and related ammonium absorption were assayed after the microalga Chlorella vulgaris Beij. was jointly immobilized with the microalgae‐growth‐promoting bacterium Azospirillum brasilense. At initial concentrations of 3, 6, and 10 mg · L^?1 NH₄⁺, joint immobilization enhances growth of C. vulgaris but does not affect ammonium absorption capacity of the microalga. However, at 8 mg · L^?1 NH₄⁺, joint immobilization enhanced ammonium absorption by the microalga without affecting the growth of the microalgal population. Correlations between absorption of ammonium per cell and per culture showed direct (negative and positive) linear correlations between these parameters and microalga populations at 3, 6, and 10 mg · L^?1 NH₄⁺, but not at 8 mg · L^?1 NH₄⁺, where the highest absorption of ammonium occurred. In all cultures, immobilized and jointly immobilized, having the four initial ammonium concentrations, enzymatic activities of Chlorella are affected by A. brasilense. Regardless of the initial concentration of ammonium, GS activity in C. vulgaris was always higher when jointly immobilized and determined on a per‐cell basis. When jointly immobilized, only at an initial concentration of 8 mg · L^?1 NH₄⁺ was GDH activity per cell higher.     

LIGHT ABSORPTION AND QUANTUM YIELD FOR GROWTH IN FIVE SPECIES OF MARINE MACROALGA1

Light utilization efficiency in five species of marine macroalgae was measured in laboratory growth experiments (13–41 days duration) at different irradiances at 7°C. All species acclimated to irradiance by changing their light absorption, resulting in a peak in light absorption between 2 and 15 μmol·m^?2.s^?1. Light absorption increased with thallus-specific chlorophyll and carbon content according to linear inverse relationships between chlorophyll content (chlarea^?1) and log[transmission] and between log[carbon content, Carea^?1] and log[transmission]. Quantum yields for light-limited growth and estimated gross photosynthesis were calculated based on incident and absorbed light. Quantum yield for photosynthesis based on light absorbed by pigments was high (mean = 114 mmol C·mol^?1 photons) and similar among the species. Quantum yield for net growth based on incident light was also high but more variable, between 22 and 75 mmol C·mol^?1 photons. Differences among species were mainly due to differences in light absorption. In conclusion, all species acclimated to low light by increasing light absorption to the maximum attainable, and growth efficiencies based on absorbed light were close to the maximum theoretically possible.     

Base Pairing at the Stem-loop Junction in the SL1 Kissing Complex of HIV-1 RNA: A Thermodynamic Study Probed by Molecular Dynamics Simulation

Abstract

The thermodynamics of the opening/closure process of a GC base pair located at the stem-loop junction of the SL1 sequence from HIV-1_Lai genomic RNA was investigated in the context of a loop-loop homodimer (or kissing complex) using molecular dynamics simulation. The free energy, enthalpy and entropy changes for the closing reaction are 0 kcal·mol^?1, ?11 kcal·mol^?1and ?0.037 kcal·mol^?1-K^?1 at 300° K respectively. Furthermore it is found that the free energy change is the same for the formation of a 11 nucleotide loop closed with UG and for the formation of a 9 nucleotide loop closed with GC. Our study evidences the high flexibility of the nucleotides at the stem-loop junction explaining the weak stability of this structure.     

Kinetic and thermodynamic studies on nitrobenzylthioinosine binding to the nucleoside transporter of chinese hamster ovary cells

The binding of [G-³H]nitrobenzylthioinosine to intact Chinese hamster ovary cells has been studied kinetically and thermodynamically. The association of nitrobenzylthioinosine with cells is a second-order process which proceeds at 24°C with a rate constant of 2·10⁷ M^?1·s^?1. Dissociation of the complex was characterized as a simple first-order process with rate constant on the order of 7·10^?3 s^?1. The quotient of these is comparable to the dissociation constant as measured in equilibrium binding studies, 2.2·10^?10 M. The temperature dependence of the rate of association indicated an Arrhenius activation energy of 8.4 kcal·mol^?1, while that of the equilibrium constant for dissociation indicated a standard enthalpy change of 8.8 kcal·mol^?1. The large increase in affinity of nitrobenzylthioinosine as compared to natural nucleosides is attributable to an entropy-driven interaction with the binding site. Thymidine, dipyridamole and papaverine each decrease the apparent dissociation constant for the nitrobenzylthioinosine-cell complex; the latter, inhibitors of nucleoside transport, decrease the rate of dissociation of the complex.     

LIGHT DEPENDENCE OF GROWTH AND PHOTOSYNTHESIS IN PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE)1

Phaeodactylum tricornutum Bohlin was maintained in exponential growth over a range of photon flux densities (PFD) from 7 to 230 μmol·m^?2s^?1. The chlorophyll a-specific light absorption coefficient, maximum quantum yield of photosynthesis, and C:N atom ratio were all independent of the PFD to which cells were acclimated. Carbon- and cell-specific, light-satuated, gross photosynthesis rates and dark respiration rates were largely independent of acclimation PFD. Decreases in the chlorophyll a-specific, gross photosynthesis rate and the carbon: chlorophyll ratio and increases of cell- or carbon-specific absorption coefficients were associated with an increase in cell chlorophyll a in cultures acclimated to low PFDs. The compensation PFD for growth was calculated to be 0.5 μmol·m^?2s^?1. The maintenance metabolic rate (2 × 10^?7s^?1), calculated on the basis of the compensation PFD, is an order of magnitude lower than the measured dark respiration rate(2.7 × 10^?6mol O₂·mol C^?1s^?1). Maintenance of high carbon-specific, light-saturated photosynthesis rates in cells acclimated to low PFDs may allow effective use of short exposures to high PFDs in a temporally variable light environment.     

CRITICAL IRRADIANCE LEVELS AND THE INTERACTIVE EFFECTS OF QUANTUM IRRADIANCE AND DOSE ON GAMETOGENESIS IN THE GIANT KELP,MACROCYSTIS PYRIFERA1

Gametophytes of Macrocystis pyrifera (L.) C. Ag. were cultured under a series of quantum irradiances in three photoperiod regimes. The quantum irradiances in each photoperiod were adjusted to provide equal daily irradiation dosages between photoperiods which allowed a critical examination of the interactions between quantum irradiance and quantum dose in determining gametophyte fertility. The lowest quantum irradiance which stimulated gametogenesis in more than 50% of the female gametophytes was 5 μE·m^?2·s^?1. The saturating irradiance was ca. 10 μE·m^?2·s^?1 at photoperiods of 12 h or greater. In terms of daily quantum dose, the lowest dose at which greater than 50% gametogenesis occurred was 0.2 E·m^?2·d^?1. However, this critical quantum dose was higher (0.4 E·m^?2·d^?1) when instantaneous irradiances were less than 5 μE·m^?2·s^?1. The saturation quantum dose was also affected by the rate at which the quantum dose was received and varied from 0.4 to 0.8 E·m^?2·d^?1. Gametophytes in all three photoperiods reached 100% fertility at quantum irradiances above 5 μE·m^?2·s^?1. Photoperiod effects were small and could be accounted for by quantum dosage effects.     

Interaction of elongation factor EF-Tu with γ-amides of GTP and β-amides of GDP bearing the azidoaryl group or the chloroethylaminoaryl group placed at the terminal phosphate

New types of azidoaryl analogs of GTP: γ-(4-azido)anilide of GTP (I), γ-(N-(4-azidobenzyl)-N-methyl)amide of GTP (II) and of GDP: β-(4-azido)anilide of GDP (III), β-(N-(4-azidobenzyl)-N-methyl)amide of GDP (IV) have been synthesized by treatment of the nucleotide in aqueous solution with N-cyclohexyl-N′-β-(4-methylmorpholinium)- ethylcarbodiimidep-toluene sulfonate and the respective amine. The analog of GTP bearing at the γ-phosphate an alkylating 2-chloroethylamino group: γ-(4-N-(2-chloroethyl)-N-methylaminobenzyl)amide of GTP (V) was prepared by the method described previously for the preparation of the analog of ATP (Knorre, D.G., Kurbatov, V.A. and Samukov, V.V. (1976) FEBS Lett. 70, 105–108). Azidoaryl analogs of GTP and GDP as well as the chloroethylaminoaryl analog of GTP compete with GDP in the formation of the binary complex EF-Tu·GDP with the respective K_i values 3.9·10^?7 M (I), 2.9·10^?8 M (II), 6.9·10^?7 M (III), 5.0·10^?7 M (IV) and 3.8·10^?8 M (V) relative to GDP. constants of the complexes of the radioactively-labeled GTP analogs I, II and V with elongation factor Tu were calculated to be 8.5·10^?6 M, 3.4·10^?7 M and 4.6·10^?8 M, respectively, or approx. 1740-, 70- and 9-times greater than that of GDP. GTP analogs I, II and V were found to substitute GTP in the stimulation of EF-Tu-dependent binding of aminoacyl-tRNA to the ribosome-mRNA complex.     

Synthesis,characterization, DNA interaction,and cytotoxicity of novel Pd(II) and Pt(II) complexes

Four complexes [Pd(L)(bipy)Cl]·4H₂O (1), [Pd(L)(phen)Cl]·4H₂O (2), [Pt(L)(bipy)Cl]·4H₂O (3), and [Pt(L)(phen)Cl]·4H₂O (4), where L = quinolinic acid, bipy = 2,2’-bipyridyl, and phen = 1,10-phenanthroline, have been synthesized and characterized using IR, ¹H NMR, elemental analysis, and single-crystal X-ray diffractometry. The binding of the complexes to FS-DNA was investigated by electronic absorption titration and fluorescence spectroscopy. The results indicate that the complexes bind to FS-DNA in an intercalative mode and the intrinsic binding constants K of the title complexes with FS-DNA are about 3.5?×?10⁴ M^?1, 3.9?×?10⁴ M^?1, 6.1?×?10⁴ M^?1, and 1.4?×?10⁵ M^?1, respectively. Also, the four complexes bind to DNA with different binding affinities, in descending order: complex 4, complex 3, complex 2, complex 1. Gel electrophoresis assay demonstrated the ability of the Pt(II) complexes to cleave pBR322 plasmid DNA.     

Kocuria SM1 controls vibriosis in rainbow trout (Oncorhynchus mykiss,Walbaum)

Aims: To develop probiotics for the control of vibriosis caused by Vibrio anguillarum and Vibrio ordalii in finfish. Methods and Results: Kocuria SM1, isolated from the digestive tract of rainbow trout, was administered orally to rainbow trout (Oncorhynchus mykiss) for 2 weeks at a dose equivalent to c. 10⁸ cells per g of feed and then challenged intraperitoneally with V. anguillarum and V. ordalii. Use of SM1 led to a reduction in mortalities to 15–20% compared to 74–80% mortalities in the controls. SM1 stimulated both cellular and humoral immune responses in rainbow trout, by elevation of leucocytes (5·5 ± 0·8 × 10⁶ ml^?1 from 3·7 ± 0·8 × 10⁶ ml^?1), erythrocytes (1·2 ± 0·1 × 10⁸ ml^?1 from 0·8 ± 0·1 × 10⁸ ml^?1), protein (23 ± 4·4 mg ml^?1 from 16 ± 1·3 mg ml^?1), globulin (15·7 ± 0·2 mg ml^?1 from 9·9 ± 0·1 mg ml^?1) and albumin (7·3 ± 0·2 mg ml^?1 from 6·1 ± 0·1 mg ml^?1) levels, upregulation of respiratory burst (0·05 ± 0·01 from 0·02 ± 0·01), complement (56 ± 7·2 units ml^?1 from 40 ± 8·0 units ml^?1), lysozyme (920 ± 128·8 units ml^?1 from 760 ± 115·3 units ml^?1) and bacterial killing activities. Conclusions: Kocuria SM1 successfully controlled vibriosis in rainbow trout, and the mode of action reflected stimulation of the host innate immune system. Significance and Impact of the Study: Probiotics can contribute a significant role in fish disease control strategies, and their use may replace some of the inhibitory chemicals currently used in fish farms.