Purification and characterization of thermostable β-mannanase from a Bacillus sp. YA-14

Thermostable β-mannanase from Bacillus sp. YA-14 was purified by acetone precipitation, CM-cellulose, Sephadex G-100 and hydroxyapatite column chromatography from culture supernatant. The final enzyme preparation appeared to be homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). β-Mannanase appeared to be a monomeric protein with a molecular weight of 67,000 daltons. The optimal pH and temperature of the enzyme reaction were pH 6.0 and 75°C, respectively. The enzyme was stable at a pH range of 6.0 to 9.0 and at temperatures between 45 and 85°C. The kinetic constants of β-mannanase as determined with a galactomannan (locust bean) as substrate were a Vmax of 25 unit/ml and a Km of 1.1 mg/ml. The enzyme had only limited activity on galactomannan substrate. It was suggested that β-mannanase activity is limited by the number of branched α-galactose residues.