Thermostable β-glucosidase with a broad substrate specifity suitable for processing of lactose-containing products

Sulfolobus shibatae produces intracellular β-glucosidase with β-galactosidase activity. The final yield of the wet cells after 72 h of growth at 75 °C (pH 3.5) was about 3.4 g per litre of the medium. Addition of 1% of lactose to the medium enhanced the enzyme activity by 40%. The cell-free extract of Sulfolobus shibatae had specific activity towards GlcβpNp (0.81 U mg−1), GalβoNp (1.85 U mg−1), cellobiose (0.47 U mg−1) and lactose (0.29 U mg−1). The enzyme was purified about 17-fold by ammonium sulphate precipitation, ion-exchange chromatography and gel filtration. The obtained preparation exhibited highest specific activity for GalβoNp hydrolysis (29.5 U mg−1) at pH 5.5 and 98 °C. After 5 h of preincubation at 80 °C in acetate buffer (pH 5.5), the enzyme retained 67% initial activity. The β-glucosidase and β-galactosidase activities of the enzyme was eliminated by Hg2+ and Zn2+ ions. Among thiol reagents tested, only 4-chloromercuribenzoate exhibited a strong inhibitory effect on the investigated enzyme.