CHARACTERIZATION OF TREHALASE ACTIVITIES FROM THE THERMOPHILIC FUNGUSSCYTALIDIUM-THERMOPHILUM

The thermophilic fungus Scytalidium thermophilum produced large amount s of intracellular and extracellular trehalase activity when grown on starch as the sole carbon source. The specific activity of the purifie d proteins: 1700 U (mg protein)(-1) (extracellular) and 3700 U (mg pro tein)(-1) (intracellular), was many times higher than the values repor ted for other microbial sources. The apparent molecular mass of the na tive enzymes was estimated to be 370 kDa (extracellular trehalase) and 398 kDa (intracellular trehalase) by gel-filtration chromatography. A nalysis by SDS-PAGE showed unique polypeptide bands of approx. 82 kDa (extracellular trehalase) and 85 kDa (intracellular trehalase), sugges ting that the native enzymes were composed of five subunits. The carbo hydrate content of extracellular and intracellular trehalases was esti mated to be 81% and 51%, respectively. Electrofocusing indicated a pi of 3.7 and 3.4, respectively, for the extracellular and intracellular enzymes. Both trehalases were highly specific for trehalose and were s timulated by calcium and manganese. Calcium and manganese also protect ed both trehalases from thermoinactivation. Inhibition was observed in the presence of aluminium, mercurium, copper, zinc, EDTA, ADP, and AT P. Apparent K-m values, for the extracellular and intracellular trehal ases, were 3.58 mM and 2.24 mM, respectively. The optimum of pH for th e extracellular and the intracellular trehalase was 6.0, and the optim um of temperature 60 degrees C and 65 degrees C, respectively.