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Updated: Dec 12, 2025
Efficient Mammalian Cell Expression and Single-step Purification of Extracellular Glycoproteins for Crystallization
Published on: December 23, 2015
A simplified protein purification method through nickel cleavage of the recombinant protein from the Escherichia coli
Shanqing Huang1, Tianbiao Wei, Wanxing Sha
1State Key Laboratory of Coordination Chemistry, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210023, China. shanqinghuang36@sina.com chenhao@nju.edu.cn.
Abstract:
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.
