Purification and properties of a basic endo-beta-1,6-glucanase (BGN16.1) from the antagonistic fungus Trichoderma harzianum

The antagonistic fungus Trichoderma harzianum CECT 2413 produces at least t wo extracellular beta-1,6-glucanases, among other hydrolases acting on poly saccharides from fungal cell walls, when grown in chitin as the sole carbon source. We have previously reported on the purification and biochemical ch aracterization of the major activity, which corresponds to an acidic enzyme named BGN16.2 [de la Cruz, J., Pintor-Toro, J.A., Benitez, T. & Llobell, A . (1995) J. Bacteriol. 177, 1864-1871]. In this paper, we report on the pur ification to electrophoretical homogeneity of BGN16.1, the second beta-1,6- glucanase enzyme. BGN16.1 was purified by ammonium sulfate precipitation fo llowed by adsorption and digestion of pustulan (a beta-1,6-glucan), chromat ofocusing and gelfiltration chromatography. BGN16.1 is a non-glycosylated p rotein with an apparent molecular mass of 51 kDa and a basic isoelectric po int (pI 7.4-7.7). The enzyme was active toward substrates containing beta-1 ,6-glycosidic linkages, including yeast cell walls. The K-m was 0.8 mg.mL(- 1) with pustulan as the substrate. Reaction product analysis by HPLC clearl y indicated that BGN16.1 has an endo-hydrolytic mode of action. The probabl e role of this enzyme in the antagonistic action of T. harzianum is also di scussed.