A simplified protein purification method through nickel cleavage of the recombinant protein from the Escherichia coli cell surface
| Autor: | Yufei Jiang, Jue Wang, Tianbiao Wei, Qingyuan Hu, Yingying Zhang, Shanqing Huang, Sha Wanxing, Hao Chen |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Chromatography Chemistry 010501 environmental sciences Cleavage (embryo) medicine.disease_cause 01 natural sciences Biochemistry Analytical Chemistry Cell membrane 03 medical and health sciences 030104 developmental biology medicine.anatomical_structure Protein purification Electrochemistry Cell disruption medicine Environmental Chemistry Centrifugation Target protein Bacterial outer membrane Escherichia coli Spectroscopy 0105 earth and related environmental sciences |
| Zdroj: | The Analyst. 145:6227-6231 |
| ISSN: | 1364-5528 0003-2654 |
| DOI: | 10.1039/d0an01060j |
| Popis: | To simplify the protein purification process, we developed a novel one-step purification method in which the recombinant protein can be cleaved directly from the Escherichia coli cell surface. This method involves fusion of the target protein to the C-terminus of a LOS tag comprising a surface anchor protein (Lpp-OmpA) and a sequence-specific nickel-assisted cleavage (SNAC)-tag. The LOS tag facilitates the anchoring of the target protein to the outer membrane of E. coli cells and its separation from the cell membrane through Ni2+ cleavage. Intact, biologically active protein with a purity of 95% and a yield of approximately 100 mg per liter of culture can be readily obtained through Ni2+ cleavage in resuspension solution followed by centrifugation. In this study, a practical and promising protein purification method has been established with minimal labor and cost, as no cell disruption and chromatographic separation are required downstream. |
| Databáze: | OpenAIRE |
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