Isolation and characterization of a thermostable endo-β-glucanase active on 1,3-1,4-β-d-glucans from the aerobic fungus talaromyces emersonii CBS 814.70

Abstract

A novel endoglucanase active on 1,3-1,4-β-d-glucans was purified to apparent homogeneity from submerged cultures of the moderately thermophilic aerobic fungus Talaromyces emersonii CBS 814.70. The enzyme is a single subunit glycoprotein with Mr and pI values of 40.7 ± 0.3 kDa and 4.4, respectively, and an estimated carbohydrate content of 77% (w/w). The purified β-glucanase displayed activity over broad ranges of pH and temperature, yielding respective optima values of pH 4.8 and 80°C. This enzyme was markedly thermostable with 15% of the original activity remaining after incubation for 15 min at 100°C. Substrate specificity studies revealed the identity of the enzyme to be a 1,3-1,4-β-d-glucanase. Identical Km values (13.38 mg.ml−1) were obtained with lichenan and BBG, while the Vmax value with lichenan (142.9 IU.mg−1) was approximately twice the value obtained with BBG (79.3 IU.mg−1). Time-course hydrolysis of barley-β-glucan did not proceed linearly with respect to time indicating an ‘endo’ or more processive action for the enzyme. HPAEC fractionation of the products of hydrolysis yielded a range of oligosaccharides, with cellobiose, cellotriose and cellotetraose being the predominant oligosaccharide products.