PURIFICATION AND PROPERTIES OF BETA-GALACTOSIDASE FROM PENICILLIUM-CITRINUM

A β-D-galactopyranosidase was purified to apparent homogeneity from aqueous extract ofwheat bran culture of a strain of Penicillium citrinum. The purified enzyme had a specific activity (ONPG as a substrate) of about 100 units per mg of protein. Molecular weight was estimated to be about 100,000 by gel filtration and about 110,000 by sedimentation equilibrium. SDS-electrophoresis gave a single protein band corresponding to a molecular weight of about 60,000, indicating that the enzyme consisted of two subunits. The enzyme was a glycoprotein containing about 6%carbohydrates. Isoelectric point was pH 6.4. Amino acid composition indicated relatively low contents of methionine and cysteine or cystine. Histidine was the amino-terminal residue. The enzyme was most active at pH 4.5 (at 40°C for 15 min) and at 50°C (at pH 4.5 for 15 min), and was stable at pH 4.5 to 7 (at 40°C for 3 hr) and below 40°C (at pH 5.6 for 15 min). Cu2 + and Hg2+ inhibited the activity and other ions tested had no effect. EDTA and PCMB did not affect the activity. NaCl, KCl, MgCl2 and MnCl2 had no effect on stability. The enzyme was activc only on β-D-galactopyranosides. The activity to ONPG was highest among galactosides tested and about 4-times higher than that to lactose. The apparent Km for ONPG and for lactose was 1.7 and 25 mM, respectively. No substrate inhibition, product inhibition or inhibition by p-aminophenyl-, β-D-thiogalactopyranoside was observed with this enzyme. © 1979 Taylor & Francis Group, LLC.