One-step purification of a recombinant beta-galactosidase using magnetic cellulose as a support: Rapid immobilization and high thermal stability

• Published in: Bioresource technology 2022-02, Vol.345, p.126497-126497, Article 126497
• Main Authors: Gennari, Adriano, Simon, Renate, Sperotto, Nathalia Denise de Moura, Bizarro, Cristiano Valim, Basso, Luiz Augusto, Machado, Pablo, Benvenutti, Edilson Valmir, Da Cas Viegas, Alexandre, Nicolodi, Sabrina, Renard, Gaby, Chies, Jocelei Maria, Volpato, Giandra, Volken de Souza, Claucia Fernanda
• Format: Article
• Language: English
• Subjects: beta-Galactosidase - metabolism; CBD; Cellulose; Enzyme Stability; Enzymes, Immobilized - metabolism; Hydrolysis; Lactase; Lactose; Lactose hydrolysis; Magnetic particles; Magnetic Phenomena
• ISSN: 0960-8524; 1873-2976
• DOI: 10.1016/j.biortech.2021.126497

[Display omitted] •One-step purification and targeted immobilization process of a β-galactosidase.•These bioprocesses were carried out efficiently on the magnetic cellulose supports.•Derivatives showed a typical response of a predominantly superparamagnetic system.•Immobilization process on magnetic supports increased the enzyme thermal stability.•The materials used in the immobilization processes did not show a cytotoxic effect. For the first time, this work reported the one-step purification and targeted immobilization process of a β-galactosidase (Gal) with the Cellulose Binding Domain (CBD) tag, by binding it to different magnetic cellulose supports. The process efficiency after β-galactosidase-CBD immobilization on magnetic cellulose-based supports showed values of approximately 90% for all evaluated enzymatic loads. Compared with free Gal, derivatives showed affinity values between β-galactosidase and the substrate 1.2 × higher in the lactose hydrolysis of milk. β-Galactosidase-CBD's oriented immobilization process on supports increased the thermal stability of the immobilized enzyme by up to 7 × . After 15 cycles of reuse, both enzyme preparations showed a relative hydrolysis percentage of 50% of lactose in milk. The oriented immobilization process developed for purifying recombinant proteins containing the CBD tag enabled the execution of both steps simultaneously and quickly and the obtention of β-galactosidases with promising catalytic characteristics for application in the food and pharmaceutical industries.