Abstract: Different members of the alcohol oxidoreductase family can transfer the hydride of NAD(P)H to either the re- or the si-face of the substrate. The enantioselectivity of transfer is very variable, even for a range of substrates reduced by the same enzyme. Exploiting quantitative isotopic (2)H NMR to measure the transfer of (2)H from NAD(P)(2)H to ethanol, a range of enantiomeric excess between 0.38 and 0.98, depending on the origin of the enzyme and the nature of the cofactor, has been determined. Critically, in no case was only (R)-[1-(2)H]ethanol or (S)-[1-(2)H]ethanol obtained. By calculating the relative energies of the active site models for hydride transfer to the re- or si-face of short-chain aldehydes by alcohol dehydrogenase from Saccharomyces cerevisiae and Lactobacillus brevis, it is shown that the differences in the energy of the systems when the substrate is positioned with the alkyl group in one or the other pocket of the active site could play a role in determining stereoselectivity. These experiments help to provide insight into structural features that influence the potential catalytic flexibility of different alcohol dehydrogenase activities.
Abstract: We report here a study on the physicochemical properties of cationic phospholipids liposomes used for lipoplex formulation and DNA transfer. The original cationic phospholipids synthesized in our laboratory are first presented with the liposome formulation process. The second part deals with the liposomes fusogenic properties studied by fluorescence resonant energy transfer (FRET). The nature of the cationic polar head and the formulation with or without a neutral colipid have a great influence on the FRET signal. The third part reports the study of the viscosity of the liposome by fluorescence anisotropy measurements. It has been observed that the vectors having a saturated lipid chain exhibit a more pronounced anisotropy than those having unsaturated lipid chains. Finally, liposomes formed by a mixture of phospholipids and DC-Chol (a rigid lipid) leads to increase the anisotropy denoting a more rigid liposome.
Abstract: The anaerobic fermentation of glucose by Leuconostoc mesenteroides via the reductive pentose phosphate pathway leads to the accumulation of lactic acid and ethanol. The isotope redistribution coefficients (a(ij)) that characterize the specific derivation of each hydrogen atom in ethanol in relation to the non-exchangeable hydrogen atoms in glucose and the medium water have been determined using quantitative (2)H NMR. First, it is confirmed that the hydrogens of the methylene group are related only to the 1 and 3 positions of glucose via the NAD(P)H pool and not to the 4 position, in contrast to ethanol produced by Saccharomyces cerevisiae. Second, it is found that the conversion factors (C(f)) for the transfer of hydrogen to the pro-S and pro-R positions of the methylene group are not equivalent: the C(f)-1-R:C(f)-1-S ratio is 2.1, whereas the C(f)-3-R:C(f)-3-S ratio is 0.8. It is shown that this non-equivalence is not determined by the stereochemistry of the terminal NADH- and NADPH-dependent alcohol dehydrogenases, but is dependent on the cofactor selectivities of the reductive and oxidative steps of the reduced nucleotide cycle.
