Cloning and characterization of an exoinulinase from Bacillus polymyxa

A gene encoding an exoinulinase (inu) from Bacillus polymyxa MGL21 was cloned and sequenced. It is composed of 1455 nucleotides, encoding a protein (485 amino acids) with a molecular mass of 55 522 Da. Inu was expressed in Escherichia coli and the His-tagged exoinulinase was purified. The purified enzyme hydrolyzed sucrose, levan and raffinose, in addition to inulin, with a sucrose/inulin ratio of 2. Inulinase activity was optimal at 35 °C and pH 7, was completely inactivated by 1 mM Ag+ or Hg2+. The Km and Vmax values for inulin hydrolysis were 0.7 mM and 2500 μM min−1 mg−1 protein. The enzyme acted on inulin via an exo-attack to produce fructose mainly.