GnRH analogues containing SV-40 virus T-antigen nuclear localization sequence

To improve the efficiency of anticancer drugs due to their delivery to intracellular targets a set of GnRH analogues containing nuclear localization signal (NLS) of SV-40 virus large T-antigen have been synthesized. NLS was attached to the parent molecule via ε-amino group of D-Lysine in position 1 or 6 of peptide sequence using orthogonal protection strategy. The biological activity studies revealed that incorporation of NLS moiety significantly increases cytotoxic activity of palmitoyl-containing GnRH analogues in vitro. The influence of tested peptides on tumor cells does not accompanied by the destruction of cell membrane, as confirmed in experiments with normal fibroblasts, used as a control.     

Isolation and properties of oxaloacetate keto-enol-tautomerases from bovine heart mitochondria

Two highly purified proteins with quite different properties capable of oxaloacetate keto-enol-tautomerase activity (oxaloacetate keto-enol-isomerase, EC 5.3.2.2) were isolated from the bovine heart mitochondrial matrix. The first protein has an apparent molecular mass of 37 kDa as determined by SDS-gel electrophoresis and Sephacryl SF-200 gel filtration. It is quite stable upon storage at 40 degrees C and reaches the maximal catalytic activity at pH 8.5 with a half-maximal activity at pH 7.0. The enzyme is specifically inhibited by oxalate and diethyloxaloacetate. When assayed in the enol----ketone direction at 25 degrees C (pH 9.0), the enzyme obeys a simple substrate saturation kinetics with Km and Vmax values of 45 microM and 74 units per mg of protein, respectively; the latter value corresponds to the turnover number of 2700 min-1. The second protein has an apparent molecular mass of 80 kDa as determined by SDS-gel electrophoresis and Sephacryl SF-300 gel filtration. The enzyme is rapidly inactivated at 40 degrees C and shows a sharp pH optimum of activity at pH 9.0. The enzyme can be completely protected from thermal inactivation by oxaloacetate and dithiothreitol. The kinetic parameters of the enzyme as assayed in the enol----ketone direction at 25 degrees C (pH 9.0) are: Km = 220 microM and Vmax = 20 units per mg of protein; the latter corresponds to the turnover number of 1600 min-1. The enzyme activity is specifically inhibited by maleate and pyrophosphate. About 30% of the total oxaloacetate tautomerase activity in crude mitochondrial matrix is represented by the 37 kDa enzyme and about 70% by the 80 kDa protein.     

Non-classic effects of luliberin confirming the pleiotropy of neuropeptides

The effects of synthetic neuropeptide LH-RH and its analogues were studied in experiments on 174 white male rats. The influence of the substances was shown on instrumental avoidance learning in Y-shaped maze. Convulsive and anticonvulsive effects of the preparations were studied on experimental model of corasol seizures. The analgetic effect of the substances was evaluated by behavioural pain reaction to electrical stimulation of the tail root. Analgetic, anticonvulsive, and psychostimulating LH-RH properties confirm polyfunctionality and "pleiotropy" of neuropeptides as a class of new endogenous informational compounds.     

Serotonin content in different sections of the brain, liver, intestines and blood of rats predisposed and not predisposed to alcohol consumption

Experiments were made with white random-bred rats (males) exposed to ethanol. The content of serotonin measured by spectrofluorometry was higher in the hypothalamus, brain stem and intestine, and was lower in the thalamus, striatum liver and blood in the animals predisposed to voluntary alcohol consumption and with lateral position duration 62 +/- 18 min as compared with the animals not predisposed to alcohol consumption and with lateral position duration 196 +/- 23 min, the dose of ethanol being 4.5 g/kg i. p. Thirty minutes after ethanol administration in a dose of 2.5 g/kg i. p. to the alcohol-predisposed rats there was a lowering of the serotonin content in the hypothalamus and an increase in the thalamus, brain stem, liver and blood. Meanwhile in the rats not predisposed to alcohol consumption, the serotonin content rose in the hypothalamus, brain stem, liver, intestine and blood and fell in the thalamus and striatum. It is assumed that the serotoninergic system of the brain may play a role in the formation of "positive" or "negative" attitudes to ethanol in the population of white random-bred rats.     

Specific Features of Accumulation Kinetics of Entomocidal Exotoxins of Bacillus thuringiensis Subsp. thuringiensisduring the Fermentation Stage of Bitoxibacillin Production

Accumulation of Bacillus thuringiensis subsp. thuringiensis -exotoxin (BET) in the course of industrial fermentation (a stage in the production of the entomocidal biopreparation bitoxibacillin) has been studied. It has been demonstrated in model experiments that the decrease in the content of BET in the culture fluid is accounted for by the toxin interaction with an attendant product, the exogenous metabolite (EM). EM has been isolated from the culture fluid and characterized. EM causes alkalization of the medium, exerts entomocidal effects (in Musca domestica), and fails to form salts on treatment with BaCl₂. The absorption spectrum of EM is similar to that of BET, showing a maximum at = 259 nm. The light-absorbing chromophore is a pyrimidine or purine base. A method for quantitative determination of both exotoxins (BET and EM) in bacterial preparations has been developed.     

New version of Monte Carlo expanded ensemble method for precise calculations of free energy difference

A new version of Monte Carlo (MC) expanded ensemble (EE) method is proposed for the calculations of free energy difference (FED) between two different systems with close values of the free energy. In order to check the method the FED between simple model systems (fluid of hard spheres and freely jointed polymer chain of hard spheres) was calculated. The free energy of the mentioned above systems was also calculated by a standard MC EE method in order to compare the results of two simulations. It was shown that the accuracy of a new algorithm is the same as of a standard one. At the same time new version of EE allows us to obtain FED between two systems having quite different structures, but similar free energies, during one simulation run.     

Biosynthesis of gold nanoparticles by Azospirillum brasilense

Plant-associated nitrogen-fixing soil bacteria Azospirillum brasilense were shown to reduce the gold of chloroauric acid to elemental gold, resulting in formation of gold nanoparticles. Extracellular phenoloxidizing enzymes (laccases and Mn peroxidases) were shown to participate in reduction of Au⁺³ (HAuCl₄) to Au⁰. Transmission electron microscopy revealed accumulation of colloidal gold nanoparticles of diverse shape in the culture liquid of A. brasilense strains Sp245 and Sp7. The size of the electron-dense nanospheres was 5 to 50 nm, and the size of nanoprisms varied from 5 to 300 nm. The tentative mechanism responsible for formation of gold nanoparticles is discussed.     

Influence of thyroliberin and its analogs with different hormonal activity on the pharmacological effects of ethanol

TRH and its two analogs with modified hormonal activity were examined for the capacity to antagonize acute and chronic effects of ethanol in mice. It has been demonstrated that L-pyroglutamyl-L-seryl-L-leucinamide, an analog of TRH, that does not affect the secretion of TSH and decreases prolactin production has the same capacity as TRH to reduce the time of ethanol narcosis but produces a lesser effect on the ethanol-induced fall of rectal temperature. Both the drugs did not affect the ethanol-altered ability of mice to hold on the rotating bar. Methyl ether of TRH, a hormonally inactive analog, was ineffective as shown by all the tests. Neither TRH nor its analogs changed the development of tolerance to chronic administration of ethanol, recorded by the rotating bar test and rectal temperature drop.