Synthesis of a library of xylogluco-oligosaccharides for active-site mapping of xyloglucan endo-transglycosylase.
Complex oligosaccharides containing alpha-D-xylosyl-(1-->6)-beta-D-glucosyl residues and unsubstituted beta-(1-->4)-linked D-glucosyl units were readily synthesized using enzymatic coupling catalyzed by the Cel7B E197A glycosynthase from Humicola insolens. Constituting this library required four key steps: (1) preparing unprotected building blocks by chemical synthesis or enzymatic degradation of xyloglucan polymers; (2) generating the donor synthon in the enzymatic coupling by temporarily introducing a lactosyl motif on the 4-OH of the terminal glucosyl units of the xylogluco-oligosaccharides; (3) synthesizing the corresponding alpha-fluorides, followed by their de-O-acetylation and the glycosynthase-catalyzed condensation of these donors onto various acceptors; and (4) enzymatically releasing lactose or galactose from the reaction product, affording the target molecules in good overall yields. These complex oligosaccharides proved useful for mapping the active site of a key enzyme in plant cell wall biosynthesis and modification: the xyloglucan endo-transglycosylase (XET). We also report some preliminary enzymatic results regarding the efficiency of these compounds.
Fauré
Régis
R
Centre de Recherche sur les Macromolécules Végétales (CERMAV-CNRS), BP53, 38041 Grenoble Cedex 9, France.
Saura-Valls
Marc
M
Brumer
Harry
H
Planas
Antoni
A
Cottaz
Sylvain
S
Driguez
Hugues
H
D001665:Binding Sites; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D020650:Combinatorial Chemistry Techniques; D016695:Glycosyltransferases / Q000737:chemistry*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis* / Q000737:chemistry; D013379:Substrate Specificity
D009844:Oligosaccharides; D016695:Glycosyltransferases; C073693:xyloglucan endotransglycosylase