Abstract
In this paper, an efficient and convenient Fe3O4/PMG/IDA-Ni2+ nanoparticles that applied to purify and immobilize his-tagged β-glucosidase was synthesized, in which, Fe3O4/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 Ni2+. 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 Fe3O4/PMG/IDA-Ni2+ nanoparticles can be considered as an excellent purification material. β-glucosidase was immobilized on the surface of Fe3O4/PMG/IDA-Ni2+ to form Fe3O4/PMG/IDA-β-glucosidase by means of covalent bound with imidazolyl and Ni2+. 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.
Highlights
Characteristics, including the small size, high surface area for the attachment of enzymes[10], superparamagnetism and low toxicity
The magnetic core-shell structured Fe3O4/PMG/Iminodiacetic acid (IDA)-Ni2+ nanoparticles were prepared, in which the core consisted of a Fe3O4 nanoparticle, and the surface of the shell was covered with abundant Ni2+
The Fe3O4 nanoparticles have an average diameter of about 200 nm (Fig. 2A), and were uniform both in shape and size
Summary
Characteristics, including the small size, high surface area for the attachment of enzymes[10], superparamagnetism and low toxicity. Most importantly, they can be separated from the reaction system. The magnetic core-shell structured Fe3O4/PMG/IDA-Ni2+ nanoparticles were prepared, in which the core consisted of a Fe3O4 nanoparticle, and the surface of the shell was covered with abundant Ni2+. BG was immobilized on their surface through combination with the imidazole group and Ni2+ to form Fe3O4/PMG/IDA-BG. The immobilization conditions, such as the added amount of BG, the incubating time and incubating temperature, were investigated. Optimal pH, temperature, thermal and storage stabilities, kinetic parameters, and reusability assay were studied
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
