Synchronized purification and immobilization of his-tagged β-glucosidase via Fe3O4/PMG core/shell magnetic nanoparticles

In this paper, an efficient and convenient Fe 3 O 4 /PMG/IDA-Ni 2+ nanoparticles that applied to purify and immobilize his-tagged β-glucosidase was synthesized, in which, Fe 3 O 4 /PMG (poly ( N, N’- methylenebisacrylamide-co-glycidyl methacrylate) core/shell microspheres were synthesized firstly using distillation-precipitation polymerization, then iminodiacetic acid (IDA) was used to open epoxy rings on the shell of microspheres to the combination of Ni 2+ . The gene of β-glucosidase that was from Coptotermes formosanus Shiraki was amplified, cloned into the expression vector pET28a with an N-terminal His-tag, and expressed in E.coli BL21. The nanoparticles showed the same purification efficiency as commercial nickel column which was a frequently used method in the field of purifying his-tagged proteins from crude cell lysates. The results indicated that Fe 3 O 4 /PMG/IDA-Ni 2+ nanoparticles can be considered as an excellent purification material. β-glucosidase was immobilized on the surface of Fe 3 O 4 /PMG/IDA-Ni 2+ to form Fe 3 O 4 /PMG/IDA-β-glucosidase by means of covalent bound with imidazolyl and Ni 2+ . The immobilized β-glucosidase exhibited excellent catalytic activity and stabilities compared with free β-glucosidase. In addition, immobilized β-glucosidase can be recycled for many times and retain more than 65% of the original activity. The materials display enormous potential in the aspect of purifying and immobilizing enzyme.