Direct purification and immobilization of his-tagged enzymes using unmodified nickel ferrite NiFe2O4 magnetic nanoparticles

Lau, E. C.H.T., Dodds, K. C., McKenna, C., Cowan, R. M., Ganin, A. Y. , Campopiano, D. J. and Yiu, H. H.P. (2023) Direct purification and immobilization of his-tagged enzymes using unmodified nickel ferrite NiFe2O4 magnetic nanoparticles. Scientific Reports, 13, 21549. (doi: 10.1038/s41598-023-48795-x) (PMID:38057439) (PMCID:PMC10700653)

Abstract

Purification of valuable engineered proteins and enzymes can be laborious, costly, and generating large amount of chemical waste. Whilst enzyme immobilization can enhance recycling and reuse of enzymes, conventional methods for immobilizing engineered enzymes from purified samples are also inefficient with multiple-step protocols, regarding both the carrier preparation and enzyme binding. Nickel ferrite magnetic nanoparticles (NiFe2O4 MNPs) offer distinct advantages in both purification and immobilization of enzymes. In this work, we demonstrate the preparation of NiFe2O4 MNPs via a one-step solvothermal synthesis and their use in direct enzyme binding from cell lysates. These NiFe2O4 MNPs have showed an average diameter of 8.9 ± 1.7 nm from TEM analysis and a magnetization at saturation (Ms) value of 53.0 emu g–1 from SQUID measurement. The nickel binding sites of the MNP surface allow direct binding of three his-tagged enzymes, D-phenylglycine aminotransferase (D-PhgAT), Halomonas elongata ω-transaminase (HeωT), and glucose dehydrogenase from Bacillus subtilis (BsGDH). It was found that the enzymatic activities of all immobilized samples directly prepared from cell lysates were comparable to those prepared from the conventional immobilization method using purified enzymes. Remarkably, D-PhgAT supported on NiFe2O4 MNPs also showed similar activity to the purified free enzyme. By comparing on both carrier preparation and enzyme immobilization protocols, use of NiFe2O4 MNPs for direct enzyme immobilization from cell lysate can significantly reduce the number of steps, time, and use of chemicals. Therefore, NiFe2O4 MNPs can offer considerable advantages for use in both enzyme immobilization and protein purification in pharmaceutical and other chemical industries.

• Item Type: Articles
• Additional Information: This work was supported by the Engineering and Physical Sciences Research Council EPSRC and Syngenta (grant number EP/L016419/1) for Ph.D. studentships to E.C.H.T.L. and C.A.McK. via the CRITICAT program. The Ph.D. studentship for R.M.C. is supported by the University of Edinburgh, CelluComp and IBioC. The Ph.D. studentship for K.C.D. is supported by the University of Edinburgh and Johnson Matthey Catalysts.
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Ganin, Dr Alexey
• Authors: Lau, E. C.H.T., Dodds, K. C., McKenna, C., Cowan, R. M., Ganin, A. Y., Campopiano, D. J., and Yiu, H. H.P.
• College/School: College of Science and Engineering > School of Chemistry
• Journal Name: Scientific Reports
• Publisher: Nature Research
• ISSN: 2045-2322
• ISSN (Online): 2045-2322
• Copyright Holders: Copyright © 2023 The Authors
• First Published: First published in Scientific Reports 13:21549
• Publisher Policy: Reproduced under a Creative Commons licence