The bacterium sp. ML52， isolated from deep-sea water， was found to synthesize an intracellular cold-adapted β-galactosidase. A novel β-galactosidase gene from strain ML52， encoding 1058 amino acids residues， was cloned and expressed in . The enzyme belongs to glycoside hydrolase family 2 and is active as a homotetrameric protein. The recombinant enzyme had maximum activity at 35 °C and pH 8 with a low thermal stability over 30 °C. The enzyme also exhibited a of 0.14 mM， a of 464.7 U/mg and a of 3688.1 S at 35 °C with 2-nitrophenyl-β-d-galactopyranoside as a substrate. Hydrolysis of lactose assay， performed using milk， indicated that over 90% lactose in milk was hydrolyzed after incubation for 5 h at 25 °C or 24 h at 4 °C and 10 °C， respectively. These properties suggest that recombinant sp. ML52 β-galactosidase is a potential biocatalyst for the lactose-reduced dairy industry.