Catalytic properties and mode of action of three endo-β-glucanases from Talaromyces emersonii on soluble β-1,4- and β-1,3;1,4-linked glucans

In this paper, we present the first detailed analysis of the modes of action of three purified, thermostable endo-β-D-glucanases (EG V-VII) against a range of soluble β-linked glucans. Studies indicated that EG V-VII, purified to homogeneity from a new source, the thermophilic fungus Talaromyces emersonii, are strict β-glucanases that exhibit maximum activity against mixed-link 1,3;1,4-β-D-glucans. Time-course hydrolysis studies of 1,4-β-D-glucan (carboxymethylcellulose; CMC), 1,3;1,4-β-D-glucan from barley (BBG) and lichenan confirmed the endo-acting nature of EG V-VII and verified preference for 1,3;1,4-β-D- glucan substrates. The results suggest that EG VI and EG VII belong to EC 3.2.1.6, as both enzymes also exhibit activity against 1,3-β-glucan (laminaran), in contrast to EG V. Although cellobiose, cellotriose and glucose were the main glucooligosaccharide products released, the range and relative amount of each product was dependent on the particular enzyme, substrate and reaction time. Kinetic constants (K_m, V_max, k _cat and k_cat/K_m) determined for EG V-VII with BBG as substrate yielded similar K_m and V_max values for EG V and EG VI. EG VII exhibited highest affinity for BBG (K_m value of 9.1mgml^-1) and the highest catalytic efficiency (k _cat/K_m of 12.63s^-1mg^-1ml).