A lichenase-like family 12 endo-(l→4)-β-glucanase from Aspergillus japonicus:: study of the substrate specificity and mode of action on β-glucans in comparison with other glycoside hydrolases

Using anion-exchange chromatography on Source 15Q followed by hydrophobic interaction chromatography on Source 15 Isopropyl, a lichenase-like endo-(1 -> 4)-beta-glucanase (BG, 28 kDa, pI 4.1) was isolated from a culture filtrate of Aspergillus japonicus. The enzyme was highly active against barley beta-glucan and lichenan (263 and 267 U/mg protein) and had much lower activity toward carboxymethylcellulose (3.9 U/mg). The mode of action of the BG on barley beta-glucan and lichenan was studied in comparison with that of Bacillus subtilis lichenase and endo-(1 -> 4)-beta-glucanases (EG 1, 11, and 111) of Trichoderma reesei. The BG behaved very similar to the bacterial lichenase, except the tri- and tetrasaccharides formed as the end products of beta-glucan hydrolysis with the BG contained the beta-(1 -> 3)-glucoside linkage at the non-reducing end, while the lichenase-derived oligosaccharides had the beta-(1 -> 3)linkage at the reducing end. The BG was characterized by a high amino acid sequence identity to the EG of Aspergillus kawachii (UniProt entry Q12679) from a family 12 of glycoside hydrolases (96% in 162 identified aa residues out of total 223 residues) and also showed lower sequence similarity to the EglA of Aspergillus niger (074705). (c) 2005 Elsevier Ltd. All rights reserved.