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Accueil > Publications > Recherche par années > Années 1990 > 1996

1996

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A kissing complex together with a stable dimer is involved in the HIV-1(Lai) RNA dimerization process in vitro

Retroviruses contain a dimeric RNA consisting of two identical molecules of genomic RNA. The interaction between the two monomers is thought to occur near their 5’ ends. We previously identified a region upstream from the splice donor site, comprising an autocomplementary sequence, responsible for the formation of dimeric HIV-1(Lai) RNA [Muriaux, D., Girard, P.-M., Bonnet-Mathoniere, B., & Paoletti, J. (1995) J. Biol. Chem. 270, 8209-8216]. This region appeared to be confined within a putative stem-loop structure. Here we report an in vitro model of the HIV-1 RNA dimerization process involving a two-step mechanism. We used RNA 77-402, a transcript of the HTV-1(Lai) region, which is able to dimerize spontaneously in vitro under conditions of low ionic strength.

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A model of PSI dimerization : Destabilization of the C-278-G(303) stem-loop by the nucleocapsid protein (NCp10) of MoMuLV

We have shown that at low ionic strength (i.e., 100 mM NaCl) a short autocomplementary sequence spanning nucleotides C-283 to G(298) Of MoMuLV RNA genome is involved in the process of PSI dimerization in vitro [Girard, P.-M., Bonnet-Mathoniere, B., Muriaux, D., & Paoletti, J. (1995) Biochemistry 34, 9785-9794]. In order to identify other contributions of the PSI structure to RNA dimerization, we studied the kinetics of dimerization as a function of salt concentration of short RNA transcripts comprising or not the autocomplementary sequence C-283-G(298) We propose that, apart from the crucial role of this sequence in RNA dimerization, the 364-565 domain of PSI can interfere, in vitro, with the initiation of dimer formation.

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A one-dimensional (proton and phosphorus) and two-dimensional (proton) in vivo NMR spectroscopic study of reversible global cerebral ischemia

The suitability of two-dimensional (2D) proton spectroscopy for monitoring, in vivo, the changes in levels of brain metabolites induced by cerebral ischemia was investigated in an experimental model of 30-min reversible ischemia induced by four-vessel occlusion in the rat. The resulting data were compared with those obtained by one-dimensional (1D) proton and phosphorus spectroscopy. Phosphorus spectra obtained during ischemia showed significant drops in levels of phosphocreatine (-73%), ß-ATP (-60%), and intracellular pH (to 6.30) and an increase in inorganic phosphate level (905%), 1D and 2D proton spectra showed decreases in the N-acetylaspartate/creatine-phosphocreatine ratio that were not significantly different [-21% (1D) and -32% (2D)]. Similarly, the increases in lactate/creatine-phosphocreatine ratio were not significantly different [2,546% (1D) and 3,020% (2D)].

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A short autocomplementary sequence plays an essential role in avian sarcoma-leukosis virus RNA dimerization

Retroviral genomes consist of two identical RNA molecules joined noncovalently near their 5’-ends. Recently, two models have been proposed for RNA dimer formation on the basis of results obtained in vitro with human immunodeficiency virus type 1 RNA and Moloney murine leukemia virus RNA. It was first proposed that viral RNA dimerizes by forming an interstrand quadruple helix with purine tetrads. The second model postulates that RNA dimerization is initiated by a loop-loop interaction between the two RNA molecules. In order to better characterize the dimerization process of retroviral genomic RNA, we analyzed the in vitro dimerization of avian sarcoma-leukosis virus (ASLV) RNA using different transcripts.

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Acetogenesis from H-2 and CO2 by methane- and non-methane-producing human colonic bacterial communities

The purpose of the study was to define the potential for reductive acetogenesis of colonic microflora from six non-methane- and four methane-excreting human subjects in relation to numbers of the different H-2-utilizing microorganisms. Faecal bacterial suspensions were incubated in the presence of (NaHCO3)-C-13, and under a gas phase composed of either 100% N-2 (control) or 80% H-2-20% N-2. The effects of a specific methanogenesis inhibitor or of sulfate supplementation were also determined. Quantitative nuclear magnetic resonance showed the presence of both single- and double-labelled acetate in all incubations under hydrogen. H-2/CO2-acetogenesis appears to be a quantitatively important activity only in the presence of very low numbers of methanogens. Inhibition of methanogenesis induced a large increase in (CO2)-C-13 incorporation into acetate in CH4-producing samples. These results showed that methanogens can efficiently outcompete acetogens in human colonic contents. In contrast, no clear-cut competition for H-2 between acetogenesis and dissimilatory sulfate-reduction could be demonstrated. A slight reduction of the acetogenic activity was only observed at the highest sulfate addition (100 mM).

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Carbon metabolism in Eubacterium limosum : A C-13 NMR study

Carbon metabolism was studied in Eubacterium limosum, an anaerobic acetogenic bacterium. Stationary cells fermented glucose C-13(1) to acetate-2-C-13 and butyrate-2-C-13 or butyrate-4-C-13. Reductive acetogenesis was demonstrated by feeding cells with (NaHCO3)-C-13. Acetate alone was synthesized, and the doubly labelled isotopomer was the dominant product. The biosynthesis pathways for amino acids were examined, using the incorporation of C-13. With glucose-1-C-13 as substrate, the label was found mainly on the methyl groups whereas C-13 from carbonate is incorporated mainly on carbonyl groups. The pathways operating in Eubacterium limosum were delineated with reference to the biochemistry of other microbes. It was found that Eubacterium limosum incorporates carbon through acetyl-CoA and pyruvate and the reductive TCA cycle. (C) 1996 Academic Press

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