The cloning, purification and characterisation of a cold-active β-galactosidase from the psychrotolerant Antarctic bacterium Arthrobacter sp. C2-2

The Gram-positive Antarctic bacterium Arthrobacter sp. C2-2 contains two, possibly three cold-active isoenzymes of β-galactosidase. The C2-2-1 isoenzyme was cloned, purified and characterised. This β-galactosidase was classified as being a member of the Family 2 of glycosidases. It is a homotetrameric enzyme, each subunit being composed of 1023 amino acids and it shows great activity towards lactose as a substrate. The C2-2-1 isoenzyme is particularly cold-active, compared to other β-galactosidases including those from some closely-related bacteria, retaining 20% of activity at 10 °C compared with maximum values. The temperature optimum of the purified enzyme was 40 °C using lactose as the substrate. The enzyme is particularly thermolabile, losing all activity within 10 min at 50 °C. The isoelectric point of the enzyme was 5.9. Dithiothreitol and Mg2+ ions were strong activators, whereas Cu2+, Al3+ and Tris were strong inhibitors of activity. The enzyme exhibited transglycosylation ability and the highest concentration of trisaccharides (34 mM) was formed after 10 h at 15 °C, which is an activity comparable with that of the commercial enzymes. Therefore, the C2-2-1 β-galactosidase isoenzyme of Arthrobacter sp. C2-2 has the particular advantage that it could be used as a biotechnological tool in the production of lactose-reduced dairy products at refrigeration temperatures.