BETA-1,3-GLUCAN HYDROLASES FROM EUGLENA GRACILIS .2. PURIFICATION AND PROPERTIES OF BETA-I,3-GLUCAN EXO-HYDROLASE

1. 1. The β-1,3-glucan exo- and endo-hydrolases from extracts of Euglena gracilis may be partially resolved by chromatography on Biogel P-200 and P-300. Both the exo- and endo-hydrolases are adsorbed by CM-cellulose but only the exo-hydrolase is eluted by 150 mM sodium acetate. 2. 2. The exo-hydrolase in extracts prepared at pH 4.5 and free of endo-hydrolase was chromatographed on CM-cellulose, yielding two active components. 3. 3. The first of these components was further investigated and shown to be highly specific for β 1,3-glucan substrates. Its relative rate of reaction decreased as the chain length of the substrate decreased. Laminaribiose was not hydrlysed. The Km for laminaripentaose was 11.1·10-5 M and for insoluble laminarin 0.008% (w/v). The pH-activity optimum was in the range 4.7-5.2 and the enzyme was inactive below pH-3 and above pH7. No transglycosylation from laminarin to glucose or reversion reaction with glucose alone was observed. Glucose was released in the α-configuration during hydrolysis. The enzyme was not stimulated by Mn2+ or inhibited by 1,5-gluconolactone. © 1969.