Adsorption‐based immobilization of C aldicellulosiruptor saccharolyticus cellobiose 2‐epimerase on B acillus subtilis spores

Nonrecombinant spore was examined as a novel immobilization support to adsorb enzymes. C aldicellulosiruptor saccharolyticus cellobiose 2‐epimerase ( C s CE ), efficiently producing lactulose using lactose as a single substrate, was immobilized on B acillus subtilis spores via adsorption. The immobilization process was optimized, and the properties of immobilized C s CE and the interactions between the enzyme and spores were also investigated. Under the optimized conditions (pH 4.5, temperature 4 °C, reaction time 2 H, and initial enzyme concentration 2.4 mg/mL), the maximum adsorbed amount of C s CE was 1.47 mg/10 11 spores, and the enzyme activity recovery was 79.4%. The spore‐immobilized C s CE presented a higher pH and thermal stability than a free enzyme. Total desorption of the immobilized enzyme was only achieved by treatment with 1.0 M NaCl at pH 1.0, indicating a strong adsorption between C s CE and B . subtilis spores. Efficient binding may require a potent combination of electrostatic and hydrophobic interactions between spores and an enzyme. The immobilized C s CE was applied to produce 395 g/L lactulose after 4 H. Moreover, the spores could be regenerated and the spore‐immobilized enzyme showed good reusability as it retained approximately 70% of its initial activity after eight recycles.