Conditions of formation, purification, and characterization of an α- galactosidase of Trichoderma reesei RUT C-30

Abstract

Trichoderma reesei RUT C-30 formed an extracellular α-galactosidase when it was grown in a batch culture containing lactose or locust bean gum as a carbon source. Short-chain α-galactosides (melibiose, raffinose, stachyose), as well as the monosaccharides galactose, dulcitol, arabinose, and arabitol, also induced α-galactosidase activity both when they were used as carbon sources (at a concentration of 1%) in batch cultures and in resting mycelia (at concentrations in the millimolar range). The addition of 50 mM glucose did not affect the induction of α-galactosidase formation by galactose. α- Galactosidase from T. reesei RUT C-30 was purified to homogeneity from culture fluids of galactose-induced mycelia. The active enzyme was a 50 ± 3- kDa, nonglycosylated monomer which had an isoelectric point of 5.2. It was active against several α-galactosides (p-nitrophenyl-α-D-galactoside, melibiose, raffinose, and stachyose) and galactomannan (locust bean gum) and was inhibited by the product galactose. It released galactose from locust bean gum and exhibited synergism with T. reesei β-mannanase. Its activity was optimal at pH 4, and it displayed broad pH stability (pH 4 to 8). Its temperature stability was moderate (60 min at 50°C resulted in recovery of 70% of activity), and its highest level of activity occurred at 60°C. Its action on galactomannan was increased by the presence of β-mannanase.