Aglycone specificity of Escherichia coliα-xylosidase investigated by transxylosylation.

The specificity of the aglycone-binding site of Escherichia coliα-xylosidase (YicI), which belongs to glycoside hydrolase family 31, was characterized by examining the enzyme's transxylosylation-catalyzing property. Acceptor specificity and regioselectivity were investigated using various sugars as acceptor substrates and α-xylosyl fluoride as the donor substrate. Comparison of the rate of formation of the glycosyl–enzyme intermediate and the transfer product yield using various acceptor substrates showed that glucose is the best complementary acceptor at the aglycone-binding site. YicI preferred aldopyranosyl sugars with an equatorial 4-OH as the acceptor substrate, such as glucose, mannose, and allose, resulting in transfer products. This observation suggests that 4-OH in the acceptor sugar ring made an essential contribution to transxylosylation catalysis. Fructose was also acceptable in the aglycone-binding site, producing two regioisomer transfer products. The percentage yields of transxylosylation products from glucose, mannose, fructose, and allose were 57, 44, 27, and 21%, respectively. The disaccharide transfer products formed by YicI, α-d-Xyl p-(1→6)-d-Man p, α-d-Xyl p-(1→6)-d-Fru f, and α-d-Xyl p-(1→3)-d-Fru p, are novel oligosaccharides that have not been reported previously. In the transxylosylation to cello-oligosaccharides, YicI transferred a xylosyl moiety exclusively to a nonreducing terminal glucose residue by α-1,6-xylosidic linkages. Of the transxylosylation products, α-d-Xyl p-(1→6)-d-Man p and α-d-Xyl p-(1→6)-d-Fru f inhibited intestinal α-glucosidases. [ABSTRACT FROM AUTHOR]