PARTIAL PURIFICATION AND CHARACTERIZATION OF NEUTRAL TREHALASE FROM COMMERCIAL BAKER'S YEAST, SACCHAROMYCES CEREVISIAE

Abstract

The neutral trehalase of a commercial baker's yeast (S. cerevisiae) strain has been partially purified using ammonium sulfate fractionation and DEAE-cellulose column chromatography techniques. Trehalase was precipitated between 35-50% ammonium sulfate saturation and approximately 5-8 fold purification was achieved. The yeast cAMP-dependent protein kinase was also precipitated in the same fraction and these two proteins were separated by DEAE-cellulose column chromatography. Trehalase became totally inactive after ion exchange chromatography, cryptic trehalase (tre-c), but was later activated with the addition of partially purified protein kinase together with cAMP and ATP. A 215 fold purification was obtained after DEAE-cellulose column chromatography. One mMEDTA caused complete inhibition of the enzyme in crude extract, however the inhibition levels in ammonium sulfate and DEAE-cellulose fractions were 73.5% and 50%, respectively. Optimal pH range and temperature of the enzyme were determined as pH 6-6.8 and 30C, respectively. The kinetic parameters, K m and V max , were estimated as 11.78 mM trehalose and 12.47 μmole glucose/ min-mg protein, respectively.