Site-Directed Immobilization of An Engineered Cytidine Deaminase on a Magnetic Carrier with Bacteriophage-Aided Nanosurface for Multiround Biocatalysis

Enzymes are widely applied biocatalysts. Enzyme immobilization is an effective approach to improve enzyme stability, reusability, and condition tolerance for better industrial utilization. The key factors in enzyme immobilization include versatility of the immobilization strategy, cost of the immobilization material and procedure, and catalytic performance of the immobilized enzyme. In this study, a novel composite material, a magnetic carrier with bacteriophage-aided nanosurface (McBans), has been constructed, which has combined the good features of the inorganic core of magnetic particles and the biological and nanolayer of the bacteriophage T4 capsids on the surface. By utilizing McBans for the immobilization of an engineered cytidine deaminase CD1.3, it shows well-maintained activity with an expressed activity of 89.6%, improved stability, and easy recovery of the immobilized enzyme. Enzyme purification is simultaneously achieved during immobilization with an immobilization yield of 65.1%. The immobilized CD1.3 exhibits greatly improved temperature/pH tolerance and longer shelf life and can be fully recovered after 10 reaction cycles, suggesting great advantages and application values in the biocatalysis of N-hydroxycytidine. Moreover, the versatility of McBans has been confirmed by the immobilization of an additional enzyme, an engineered l-threonine aldolase BnLTA. Overall, McBans provides a valuable enzyme immobilization tool for easier utilization of various enzymes in important enzyme-based industrial applications.