Enzymatic synthesis of N-acetylglucosaminobioses by reverse hydrolysis: characterisation and application of the library of fungal β-N-acetylhexosaminidases

The regioselectivity of 20 extracellular β-N-acetylhexosaminidases of fungal origin was screened in the reverse hydrolysis with 2-acetamido-2-deoxy-d-glucopyranose. Most of the enzymes used yielded 2-acetamido-2-deoxy-β-d-glucopyranosyl-(1→4)-2-acetamido-2-deoxy-d-glucopyranose (3) and 2-acetamido-2-deoxy-β-d-glucopyranosyl-(1→6)-2-acetamido-2-deoxy-d-glucopyranose (4). So far unknown product of enzymatic condensation, 2-acetamido-2-deoxy-β-d-glucopyranosyl-(1→3)-2-acetamido-2-deoxy-d-glucopyranose (2) was synthesised using the β-N-acetylhexosaminidases from Penicillium funiculosum CCF 1994, P. funiculosum CCF 2325 and Aspergillus tamarii CCF 1665. Addition of salts ((NH4)2SO4 or MgSO4 (0.1–1.0 M)) to the reaction increased the yields and also enhanced the β-N-acetylhexosaminidase regioselectivity.