Thermostable α-galactosidase fromBacillus stearothermophilusNUB3621: cloning, sequencing and characterization

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Abstract
An α-galactosidase gene from the thermophilic bacterium Bacillus stearothermophilus NUB3621 was cloned, sequenced, expressed in Escherichia coli and the recombinant protein was purified. The Bacillus enzyme, designated AgaN, is similar to α-galactosidases of family 36 in the classification of glycosyl hydrolases. The enzyme was estimated to be a tetramer with a molecular mass of subunits 80.3 kDa. The purified AgaN is thermostable and has a temperature optimum of activity at 75°C and a half-life of inactivation of 19 h at 70°C. AgaN displays high affinity for oligomeric substrates such as melibiose and raffinose and is able to hydrolyze raffinose in the presence of 60% sucrose with high efficiency.