Cloning, expression, and characterization of a glycoside hydrolase family 50 [beta]-agarase from a marine Agarivorans isolate

The gene for a thermostable β-agarase from Agarivorans sp. JA-1 was cloned and sequenced. It comprised an open reading frame of 2,988 base pairs, which encode a protein of 109,450 daltons consisting of 995 amino acid residues. A comparison of the entire sequence showed that the enzyme has 98.8% sequence similarities to β-agarase from Vibrio sp. JT1070, indicating that it belongs to the family glycoside hydrolase (GH)-50. The gene corresponding to a mature protein of 976 amino acids was inserted and expressed in Escherichia coli. The recombinant β-agarase was purified to homogeneity. It had maximal activity at 40°C and pH 8.0 in the presence of 1 mM NaCl and 1 mM CaCl^sub 2^. The enzyme hydrolyzed agarose as well as neoagarohexaose and neoagarotetraose to yield neoagarobiose as the main product. Thus, the enzyme would be useful for the industrial production of neoagarobiose.[PUBLICATION ABSTRACT]