Purification and characterization of an endoglucanase from Aspergillus terreus highly active against barley β-glucan and xyloglucan

Abstract

An endoglucanase (1, 4-b-D glucan glucanohy- drolase, EC 3.2.1.4) which was catalytically more active and exhibited higher affinity towards barley b-glucan, xyloglu- can and lichenin as compared to carboxymethylcellulose (CMC) was purified from Aspergillus terreus strain AN1 following ion-exchange and hydrophobic interaction chro- matography and gel filtration. The purified enzyme (40-fold) that apparently lacked a cellulose-binding domain showed a specific activity of 60 lmol mg -1 protein -1 against CMC. The purified enzyme had a molecular weight of 78 and 80 KDa as indicated by sodium dodecyl sulphate-poly- acrylamide gel electrophoresis and gel filtration, respectively, and a pI of 3.5. The enzyme was optimally active at temperature 60C and pH 4.0, and was stable over a broad range of pH (3.0-5.0) at 50C. The endoglucanase activity was positively modulated in the presence of Cu 2? , Mg 2? ,C a 2? ,N a ? , DTT and mercaptoethanol. Endoglu- canase exhibited maximal turn over number (Kcat) and catalytic efficiency (Kcat/km) of 19.11 9 10 5 min -1 and 29.7 9 10 5 mM -1 min -1 against barley b-glucan as sub- strate, respectively. Hydrolysis of CMC and barley b-glucan liberated cellobiose, cellotriose, cellotetraose and detectable amount of glucose. The hydrolysis of xyloglucan, how- ever, apparently yielded positional isomers of cellobiose, cellotriose and cellotetraose as well as larger