Purification of murine oncornaviral phosphoproteins using alkyl agarose derivatives..

Methods for the purification of both murine mammary tumor (type B) and murine leukemia (type C) oncornaviral phosphoproteins are described, in which chromatography on alkyl-agarose derivatives is used as the primary fractionation step. Gel filtration or ion exchange chromatography on phosphocellulose was the only subsequent step required for the purification of the type B and type C viral proteins, respectively. The two-step protocols also resulted in the co-purification of a low molecular weight core protein from each virus. Recoveries of the viral proteins purified by this method, based on per cent contribution of individual polypeptides to total virion proteins, were 70% or greater. Radioimmunocompetition analysis of the purified murine mammary tumor virus major core protein as well as analysis of the RNA binding properties of purified low molecular weight type C virus proteins suggests that neither antigenic reactivity nor specific RNA binding characteristics are altered by the purification protocols. The availability of these procedures should aid studies on the possible function and immunochemical properties of the native murine oncornaviral phosphoproteins and may also be extended to the purification of other oncornaviral proteins.     

Amelioration of vanadium-induced testicular toxicity and adrenocortical hyperactivity by vitamin E acetate in rats

Vanadium toxicity is a challenging problem to the health professionals and a cutting-edge medical problem. Vanadium has been recognized as industrial hazards that adversely affect human and animal reproductive health. Since testicular function is exquisitely susceptible to reactive-oxygen species, the present study elucidates the possible involvement of oxidative stress in vanadium-induced testicular toxicity and the prophylactic effects of vitamin E acetate against such adverse effects of vanadium. The study also characterizes the effects of vanadium on rat adrenal steroidogenesis and determines the underlying mechanisms of testicular and adrenal interactions in response to vanadium exposure. Significantly reduced sperm count associated with decreased serum testosterone and gonadotropins level in the vanadium-injected group of rats compared to control substantially proves the ongoing damaging effects of vanadium-induced ROS on developing germ cells. This is in turn reflected in the appreciable increase in testicular lipid peroxidation level and decline in the activities of steroidogenic and antioxidant enzymes. However, oral administration of vitamin E acetate could protect testes from the toxic effects of vanadium. Vanadium also results in adrenocortical hyperactivity, as evidenced by the elevated secretion of glucocorticoids, adrenal gland hypertrophy and increased activity of adrenal Δ⁵3β-HSD. However, reversibility of these alterations in adrenocortical activities was vividly reflected after vitamin E acetate supplementation. All these studies reveal that oxidative stress is the major mechanism of health deterioration and that vanadium can act as a stressor metal causing chronic stress effects through excitation of hypothalamo-pituitary-adrenal axis. However antioxidant support by vitamin E acetate may provide significant protection.     

Molecular and morphological characterization of somatic hybrids between <Emphasis Type="Italic">Solanum tuberosum</Emphasis> L. and <Emphasis Type="Italic">S. etuberosum</Emphasis> Lindl.

Interspecific potato somatic hybrids between Solanum tuberosum L. (di)haploid C-13 and 1 endosperm balance number non-tuberous wild species S. etuberosum Lindl. were produced by protoplasts electrofusion. The objective was to transfer virus resistance from this wild species into the cultivated potatoes. Post-fusion products were cultured in VKM medium followed by regeneration of calli in MS₁₃ K medium at 20°C under a 16-h photoperiod, and regenerants were multiplied on MS medium. Twenty-one somatic hybrids were confirmed by RAPD, SSR and cytoplasm (chloroplast/mitochondria) type analysis possessing species-specific diagnostic bands of corresponding parents. Tetraploid nature of these somatic hybrids was determined through flow cytometry analysis. Somatic hybrids showed intermediate phenotypes (plant, leaves and floral morphology) to their parents in glass-house grown plants. All the somatic hybrids were male-fertile. ELISA assay of somatic hybrids after artificial inoculation of Potato virus Y (PVY) infection reveals high PVY resistance.     

NADPH production from NADP+ with a glucose dehydrogenase system involving whole cells and immobilized cells of Gluconobacter suboxydans

Summary A convenient and efficient method of NADPH production from NADP⁺ has been established using a glucose dehydrogenase system involving whole cells and immobilized cells of Gluconobacter suboxydans IFO 3172. Using airdried cells of the bacterium, the optimum conditions for NADPH production were examined, including the cell and glucose concentrations, NADP⁺ concentration, pH, buffer and reaction temperature. Under suitable conditions, the conversion ratio and the amount of NADPH accumulated reached about 100% and 73 mg/ml of the reaction mixture, respectively, after 1-h reaction. Intact cells of the bacterium also showed high NADPH production even in the reaction mixture without a surfactant. The addition of Triton X-100 to the reaction mixture and freeze-thawing treatment of intact cells enhanced the production. The NADPH production method was further improved by using cells of the bacterium immobilized by entrapment in a -carrageenan gel lattice. The immobilized cells had almost the same enzymatic properties as the air-dried cells. The conditions for the continuous production of NADPH with an immobilized cell column were also investigated. NADPH was produced in a good yield (about 95%) with this continuous process.     

Effect of tertiary amine local anesthetics on G protein-coupled receptor lateral diffusion and actin cytoskeletal reorganization

Although widely used clinically, the mechanism underlying the action of local anesthetics remains elusive. Direct interaction of anesthetics with membrane proteins and modulation of membrane physical properties by anesthetics are plausible mechanisms proposed, although a combination of these two mechanisms cannot be ruled out. In this context, the role of G protein-coupled receptors (GPCRs) in local anesthetic action is a relatively new area of research. We show here that representative tertiary amine local anesthetics induce a reduction in two-dimensional diffusion coefficient of the serotonin_1A receptor, an important neurotransmitter GPCR. The corresponding change in mobile fraction is varied, with tetracaine exhibiting the maximum reduction in mobile fraction, whereas the change in mobile fraction for other local anesthetics was not appreciable. These results are supported by quantitation of cellular F-actin, using a confocal microscopic approach previously developed by us, which showed that a pronounced increase in F-actin level was induced by tetracaine. These results provide a novel perspective on the action of local anesthetics in terms of GPCR lateral diffusion and actin cytoskeleton reorganization.     

Green synthesis and physiochemical characterization of nickel oxide nanoparticles: Interaction studies with Calf thymus DNA

One of the most promising applications of nanomaterials is that of nanobiosensors, using biomolecules such as nucleic acids as receptors. This study aimed to synthesize nickel oxide nanoparticles (NiO NPs) by an environmentally friendly green synthesis, using the extract of the herb Coriandrum sativum (coriander). The synthesized NPs were characterized using UV–Visible spectroscopy, Fourier transform infrared spectroscopy, X‐ray diffraction, X‐ray photon spectroscopy, field emission scanning electron microscopy coupled with energy dispersive spectroscopy, dynamic light scattering, zeta potential and transmission electron microscopy. All results confirmed the synthesis of pure, spherical, positively charged NiO NPs of around 95 nm in diameter with prominent hydroxyl groups attached to the surface. Furthermore, interaction studies of synthesized NiO NPs with calf thymus DNA (CT DNA) were performed using UV–Visible spectroscopy, UV–thermal melting, circular dichroism, and fluorescence spectroscopy. CT DNA served as a substitute for nucleic acid biosensors. All experimental studies indicated that the NiO NPs bound electrostatically with CT DNA. These studies may facilitate exploring the potential of NiO NP–nucleic acid conjugated materials to be used as nanobiosensors for various applications, especially in pharmacological, epidemiological, and environmental diagnostic applications, and in detection.