Previous work has shown that a region of Moloney murine leukemia virus (MoMuLV) RNA located between nucleotides 280 and 330 in the PSI region (nt 215-565) is implicated in the dimerization process. We show with a deletion from nucleotides 290-299 in PSI RNA transcripts and through an antisense oligonucleotide complementary to nucleotides 275-291 that the 283-298 region is involved in RNA dimer formation in vitro. In an attempt to further characterize the mechanism of dimer formation, a series of short RNA transcripts was synthesized which overlapps the PSI region of MoMuLV RNA.
Light scattering and P-31-NMR have been used to monitor the effect of the bee-toxin, melittin, on phosphatidylcholine (PC) bilayers of variable acyl chain length (from C-16:0 to C-20:0). Melittin interacts with all lipids provided the interaction is initiated in the lipid fluid phase. For low-to-moderate amounts of toxin (lipid-peptide molar ratios, R(i) greater than or equal to 15), the system takes the form of large spheroidal vesicles, in the fluid phase, whose radius increases from 750 Angstrom A with dipalmitoyl-PC (DPPC) to 1500 Angstrom A with diarachinoyl-PC (DAPC). These vesicles fragment into small discoids of 100-150 Angstrom A radius when the system is cooled down below T-c (the gel-to-fluid phase transition temperature).
After the demonstration that Stigmatella aurantiaca DW4 secretes an endo N-acetyl-beta-D-glucosaminidase (ENGase), acting on the di-N-acetylchitobiosyl part of N-linked glycans (S. Bourgerie, Y. Karamanos, T. Grard, and R. Julien, J, Bacteriol, 176:6170-6174, 1994), an ENGase activity having the same substrate specificity was also found to be secreted during vegetative growth of Myxococcus xanthus DK1622. The activity decreased in mutants known to secrete less protein than the wild type (Exc()), During submerged development, the activity was produced in two steps : the first increase occurred during the aggregation phase, and the second one occurred much later, during spore formation, This production was lower in developmental mutants impairing cell-cell signaling, the late mutants (csg and dsg) being the most deficient, Finally, when sporulation was obtained either by starvation in liquid shake flask culture or by glycerol induction, the activity was produced exclusively by the wild-type cells during the maturation of the coat.
Classical models for DNA triple helix formation assume the stabilization of these structures through the formation of Hoogsteen hydrogen bonds. This assumes that G-rich duplex DNA is more stable than tripler DNA. We report the results of co-migration assay, dimethyl sulfate footprint, and UV spectroscopic melting studies that reveal that at least in some cases of short (13-mer) purine(purine-pyrimidine) tripler the stability of double-stranded DNA is increased by the binding of the third strand. Under conditions which are usually considered as physiological (10 mM MgCl2, 150 mM Na+ or K+) and with a rate of heating/cooling of 1 degrees C/min, there is a good reversibility of the melting profiles which is consistent with a high rate of tripler formation. Other factors than Hoogsteen hydrogen bonds should therefore be involved in tripler stabilization. We suggest that oligonucleotides with similar properties could be efficient agents for artificial gene regulation.
A new method for analysing complex mixtures, H-1 2D n.m.r., was used to determine polyaromatics in crude gas oil mixtures. 2D NMR overcomes the lack of resolution due to crowded 1D spectra and provides structural information. In particular, TOCSY (total correlation spectroscopy) 2D n.m.r, is well suited to polyaromatics because these molecules give specific 2D fingerprints which can be easily recognized. These patterns were selected and analysed in two ways : (1) specific fingerprint recognition using a pure compound library ; (2) using the mixing time tau(m) of the TOCSY sequence. Variation of tau(m) gives a change in cross-peak intensities. The intensity variation curves are characteristic of spin systems and hence of the structures of compounds. Alkylated substitutions were also studied. The compounds were quantified. This strategy was used to analyse crude gas oils and measure the contents of separated alkylated naphthalene isomers, biphenyls, anthracene, phenanthrene and benzothiophene.
The endo-N-acetyl-beta-D-glucosaminidases (ENGase) acting on the N-N’-diacetylchitobiosyl core of N-glycosylproteins are essential reagents for the investigation of the structure and the functions of glycoproteins. These enzymes were largely studied with the aim of offering more tools with new and broader substrate specificities to the community of glycobiologists. Conversely, little attention was given to their potential role in the physiology of bacteria, even though it had been shown that ENGases are important enzymes for the physiology of animal and plant cells. In this brief review, we present the main characteristics of the bacterial ENGases and confine our discussion to biological aspects of their action in bacterial systems.
MAMMALS react to acute hypoxia with an initial augmentation and a secondary depression of the respiratory rhythm generated by brain stem neuronal networks. To investigate the cytosolic level of energy rich phosphorus metabolites during these responses, we developed P-31 nuclear magnetic resonance spectroscopy of the brain stem. Moderate hypoxia (p(a)O(2) = 40 mmHg, 2 min) caused a reversible 62 +/- 15% respiratory rhythm depression and decreased cytosolic phosphocreatine levels by 43 +/- 11% (p
With peptide analogues of their natural substrates (the glycopeptide units of nascent peptidoglycan), the DD-peptidases exhibit a strict preference for D-Ala-D-Xaa C-termini. Gly is tolerated as the C-terminal residue, but with a significantly decreased activity. These enzymes were also known to hydrolyse various ester and thiolester analogues of their natural substrates. Some thiolesters with a C-terminal leaving group that exhibited L stereochemistry were significantly hydrolysed by some of the enzymes, particularly the Actinomadura R39 DD-peptidase, but the strict specificity for a D residue in the penultimate position was fully retained. These esters and thiolesters also behave as substrates for ß-lactamases. In this case, thiolesters exhibiting L stereochemistry in the ultimate position could also be hydrolysed, mainly by the class-C and class-D enzymes. However, more surprisingly, the class-C Enterobacter cloacae P99 ß-lactamase also hydrolysed thiolesters containing an L residue in the penultimate position, sometimes with a higher efficiency than the D isomer.
A theoretical investigation of the indirect effect of radiation on the DNA macromolecule in dilute aqueous solution has been performed. A combination of the Monte Carlo method and the model of Smoluchowski together with the knowledge of rate constants for . OH radical reaction with DNA constituents and the detailed model of the DNA atomic structure allows us to calculate patterns of . OH radical attack on DNA. These patterns depend on DNA strandedness (single or double), base sequence and DNA conformation (A-, B-, Z-DNA). The comparison of the calculated patterns with the experimental data on strand breakage induced by gamma-radiolysis enables us to discuss the results obtained in term of molecular mechanisms of radiation induced damage. We can conclude : a) for B-DNA the variation in initial damage is mainly determined by the rate constant of the chemical reaction of . OH radical with different DNA components together with their specific position in the structure (sequence), and b) base radicals contribute only slightly to the formation of frank strand breaks, but significantly to the formation of alkali revealed breaks in B-DNA in dilute aqueous solution. For A-DNA only small differences in its radiolysis, as compared to the B-DNA, are predicted. On the contrary, calculations predict substantial differences in the indirect effect of radiation on the Z-form of DNA.