Enzymatically-synthesized xylo-oligosaccharides laurate esters as surfactants of interest.

Lipase-catalyzed synthesis of xylo-oligosaccharides esters from pure xylobiose, xylotriose and xylotetraose in the presence of vinyl laurate was investigated. The influence of different experimental parameters such as the loading of lipase, the reaction duration or the use of a co-solvent was studied and the reaction conditions were optimized with xylobiose. Under the best conditions, a regioselective esterification occurred to yield a monoester with the acyl chain at the OH-4 of the xylose unit at the non-reducing end. Surface-active properties of these pure xylo-oligosaccharides fatty esters have been evaluated. They display interesting surfactant activities that differ according to the degree of polymerization (DP) of the glycone moiety.

Exploring the aglycone subsite of a GH11 xylanase for the synthesis of xylosides by transglycosylation reactions.

Xylanases Tx-xyn10 and Tx-xyn11 were compared for their transxylosylation abilities in the presence of various acceptors. Tx-xyn10 exhibited a broad specificity for various acceptors, whereas xylanase Tx-xyn11 catalysed transxylosylation reactions only in presence of polyphenolic acceptors. A modelling approach was developed to study the molecular bottlenecks into the active site of the enzyme that could be responsible for this restricted specificity. The glycosyl-enzyme intermediate of Tx-xyn11 was modelled, and a rotamer of the Y78 residue was integrated. In silico mutations of some residues from the (+1) and (+2) subsites were tested for the deglycosylation step in the presence of non-polyphenolic acceptors. The results indicated that the mutant W126A was able to use aliphatic alcohols and benzyl alcohol as acceptors for transxylosylation. Experimental validation was tested by mutating the xylanase Tx-xyn11 at position W126 into alanine. The specific activity and catalytic efficiency of the W126A mutant during the hydrolysis of xylans decreased by 2-fold and 4-fold, respectively, compared to wild-type xylanase. Among tested acceptors, transxylosylation catalysed by mutant W126A was improved with benzyl alcohol leading to a 2-fold higher concentration of benzyl xylobioside, as predicted by in silico mutation. This improved transxylosylation in the presence of benzyl alcohol leading to higher synthesis of benzyl xylobioside could likely be explained by lowest steric hindrance in the aglycone subsite of the mutated xylanase. No secondary hydrolysis of benzyl xylobioside occurred for both wild-type and mutant xylanases. Finally, our results demonstrated that the modelling approach was limited and that accounting for protein dynamics can lead to improved models.

Preparation of new ß-D-xyloside- and ß-D-xylobioside-based ionic liquids through chemical and/or enzymatic reactions.

Several tetraalkylphosphonium and tetraalkylammonium salts containing xyloside- and xylobioside-based anionic moieties have been prepared. Two stereoselective routes have been developed: i) a chemical pathway in four steps from D-xylose, and ii) a chemoenzymatic pathway directly from biomass-derived xylans. These salts displayed interesting properties as ionic liquids. Their structures have been correlated to their thermal properties (melting, glass transition and decomposition temperatures).

The mechanism of inactivation of S-adenosylhomocysteine hydrolase by fluorinated analogs of 5'-methylthioadenosine.

5'-Deoxy-5'-difluoromethylthioadenosine (DFMTA) 1a and 5'-deoxy-5'-trifluoromethyl-thioadenosine (TFMTA) 1b are inhibitors of beef liver S-adenosyl-L-homocysteine hydrolase. DFMTA and TFMTA are time-dependent and irreversible inhibitors of the enzyme. Both 1a and 1b are oxidized by E-NAD+ to produce E-NADH and fluoride anion is formed in the inactivation reaction (2.2 mol of fluoride/mole of enzyme subunit and 3.1 fluoride/mole of enzyme subunit from DFMTA and TFMTA respectively). Using [8-3H]-1a or [8-3H]-1b no trace of labelled adenosine was detected during the inactivation reaction but adenine was formed. The mechanism of inhibition of S-adenosyl-L-homocysteine hydrolase by these two fluorinated nucleosides is discussed.

Mechanistic implications of cyclic ADP-ribose hydrolysis and methanolysis catalyzed by calf spleen NAD+glycohydrolase.

Calf spleen NAD+glycohydrolase, besides its well known reactions, was shown to catalyze hydrolysis and methanolysis (with retention of configuration) of cyclic ADP-ribose, indicating that this classical NADase also belongs to the class of cyclic ADP-ribose hydrolases. No formation of cyclic ADP-ribose could be detected during the hydrolysis of beta-NAD+; moreover, the kinetic parameters of cyclic ADP-ribose seem to rule out that it is a kinetically competent reaction intermediate in the conversion of NAD+ into ADP-ribose by this enzyme.