An efficient protein immobilization strategy: protein encapsulated in nano molecular cages

Abstract

BACKGROUND: To immobilize protein firmly and maintain its native conformation, a 'nano molecular cage' was designed to encapsulate bovine serum albumin (BSA) on the surface of poly(methyl methacrylate) (PMMA) magnetic microspheres by cross-linking poly(ethylene imine) (PEI) chains. RESULTS:The amount of BSA adsorbed on PMMA@PEI microspheres decreased significantly from 70.6 to 7.6 mg g −1 microsphere after desorption under high ionic strength. A similar phenomenon could also be observed for BSA adsorbed on PMMA@PEI-CB (Cibacron Blue F3GA) microspheres. In contrast, the amount of BSA encapsulated in molecular cages only decreased a little after desorption. Furthermore, conformational analysis by Fourier transform infrared spectroscopy (FTIR) showed that the contents of -helices and -sheets decreased slightly by 3.3 and 0.3% respectively on encapsulating BSA in molecular cages. Furthermore, the effects of various factors, including glutaraldehyde amount, cross-linking time and temperature, on BSA encapsulation were investigated. CONCLUSION:The results demonstrated that molecular cages could not only immobilize protein firmly but also maintain its native conformation. Therefore molecular cage encapsulation is suggested to be a potentially valuable protocol in enzyme immobilization. Attachment of CB onto magnetic microspheresCB was attached onto PMMA@PEI magnetic microspheres by nucleophilic reactions between the amino group of PEI and the chloro group of CB. Briefly, 20 mg of PMMA@PEI magnetic microspheres were added into 2 mL of CB solution (3 mg mL −1 ) in a shaker at 160 rpm and 50 ∘ C. To deposit the dye on the microspheres, 140 mg of NaCl was added into the mixture. After 30 min, 100 mg of Na 2 CO 3 was added to speed up the reaction. The reaction was carried out for 4 h. Thereafter, the resulting PMMA@PEI-CB magnetic microspheres were washed with 950 mL L −1 alcohol and distilled water several times until no CB was detected in the supernatant.