Thermostable β-Galactosidase from an Extreme Thermophile, Thermus sp. A4: Enzyme Purification and Characterization, and Gene Cloning and Sequencing

Abstract

  We purified and characterized a thermophilic β-galactosidase from Thermus sp. A4 isolated from the Atagawa hot spring (Shizuoka, Japan). The enzyme was monomeric, and its molecular mass was estimated to be 75 kDa by SDS-polyacrylamide gel electrophoresis. The enzyme was extremely thermostable and retained its full activity after incubation at 70°C for 20 h. The K_m observed were 5.9 mM for o-nitrophenyl β-D-galactopyranoside and 19 mM for lactose. We cloned and analyzed the complete sequence of the gene encoding this enzyme. It was found to consist of 645 amino acid residues. We propose that this enzyme and seven other unclassified β-galactosidases are new members of family 42 of the glycosyl hydrolases.