Characterization of galactose-induced extracellular and intracellular pectolytic activities from the exo-1 mutant strain of Neurospora crassa

Pectolytic enzymes from the hyperproducer exo-1 mutant of Neurospora crassa are induced either by pectin or galactose. Galactose-induced pectinases, in contrast with pectin-induced enzymes, are not affected by glucose repression. Here, the pectolytic enzymes induced by galactose were purified and characterized. Extracellular pectolytic activities were separated into two main fractions. Pool I contained lyases, and a polygalacturonase (PG) copurifying as a complex of about 80 kDa (gel filtration). Pool II contained PG only. Under urea-SDS-PAGE the lyases and polygalacturonase from pool I migrated with an apparent MW of 56.2 kDa, and 34.3 kDa, respectively. PG from pool II exhibited an apparent MW of 44.7 kDa. Cell extracts contained PG free of lyase activities. Purified intracellular PG migrated (SDS-PAGE) as a single band of apparent MW of 31.5 kDa. All pectinases were glycoproteins (18.5–39% carbohydrate), with stability and optimum pH at 5–6 and 9–10 for PG and lyases, respectively. Temperature optima were 40–50°C, respectively. All enzymes were inactivated at 60°C, with a half-life from 1.5 to 5 min. Activation energy (Ea) values for extracellular and intracellular PG varied between 0.45 and 2.0 Kcal mol⁻¹. Pool II and intracellular PG and lyases, exhibited a random mechanism of hydrolysis. Pool I PG exhibited an exo character.