Please use this identifier to cite or link to this item: https://hdl.handle.net/10356/83191
Title: Going native: Complete removal of protein purification affinity tags by simple modification of existing tags and proteases
Authors: Goh, Hui Chin
Sobota, Radoslaw M.
Ghadessy, Farid J.
Nirantar, Saurabh
Keywords: Protein purification
Affinity tags
Protease
Issue Date: 2017
Source: Goh, H. C., Sobota, R. M., Ghadessy, F. J.,& Nirantar, S. (2017). Going native: Complete removal of protein purification affinity tags by simple modification of existing tags and proteases. Protein Expression and Purification, 12918-24.
Series/Report no.: Protein Expression and Purification
Abstract: Protein purification typically involves expressing a recombinant gene comprising a target protein fused to a suitable affinity tag. After purification, it is often desirable to remove the affinity tag to prevent interference with downstream functions of the target protein. This is mainly accomplished by placing a protease site between the tag and the target protein. Typically, a small oligopeptide ‘stub’ C-terminal to the cleavage site remains attached to the target protein due to the requirements of sequence-specific proteases. Furthermore, steric hindrance can also limit protease efficiency. Here, we show that respectively fusing the interacting ePDZ-b/ARVCF protein-peptide pair to the target protein and a protease enables efficient processing of a minimised sequence comprising only residues N-terminal to the cleavage site. Interaction of the protein-peptide pair enforces proximity of the protease and its minimised cleavage sequence, enhancing both catalysis of a sub-optimal site and overcoming steric hindrance. This facilitates the high yield purification of fully native target proteins without recourse to specialised purification columns.
URI: https://hdl.handle.net/10356/83191
http://hdl.handle.net/10220/42481
ISSN: 10465928
DOI: 10.1016/j.pep.2016.09.001
Schools: School of Biological Sciences 
Rights: © 2016 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Fulltext Permission: open
Fulltext Availability: With Fulltext
Appears in Collections:SBS Journal Articles

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