Stereochemistry of the hydrolysis of glycosidic linkage by endo-β-1,4-xylanases of Trichoderma reesei

Abstract

Methyl β- d-xylotrioside was used as a non-reducing substrate to investigate the stereochemistry of hydrolysis of, β-1,4-xylopyranosidic linkage by purified endo-β-1,4-xylanases (EC 3.2.1.8) of Trichoderma reesei, employing 1H NMR spectroscopy. The fungus produces one acidic species (pI 4.8–5.5), designated as EXI, and one alkaline species (pI 8.5–9.0), designated as EXII. Both enzymes were found to cleave the xylotrioside predominantly to methyl β- d-xyloside and xylobiose. Monitoring of the intensity of the H-1 signals of α- and β-xylobiose during the time course of hydrolysis clearly showed that both enzymes liberate the β-anomer of xylobiose, i.e. a product with anomeric configuration identical with that of the cleaved glycosidic linkage. This means that both EXI and EXII belong to the so-called retaining glycanases that utilize the double displacement reaction mechanism of hydrolysis.