Escherichia coli protein expression system for Acetylcholine Binding Proteins (AChBPs)

Abraham, Nikita, Paul, Blessy, Ragnarsson, Lotten and Lewis, Richard J. (2016) Escherichia coli protein expression system for Acetylcholine Binding Proteins (AChBPs). PLoS One, 11 6: . doi:10.1371/journal.pone.0157363


Author Abraham, Nikita
Paul, Blessy
Ragnarsson, Lotten
Lewis, Richard J.
Title Escherichia coli protein expression system for Acetylcholine Binding Proteins (AChBPs)
Formatted title
Escherichia coli protein expression system for Acetylcholine Binding Proteins (AChBPs)
Journal name PLoS One   Check publisher's open access policy
ISSN 1932-6203
Publication date 2016-06-15
Year available 2016
Sub-type Article (original research)
DOI 10.1371/journal.pone.0157363
Open Access Status DOI
Volume 11
Issue 6
Total pages 15
Place of publication San Francisco, United States
Publisher Public Library of Science
Collection year 2017
Language eng
Formatted abstract
Nicotinic acetylcholine receptors (nAChR) are ligand gated ion channels, identified as therapeutic targets for a range of human diseases. Drug design for nAChR related disorders is increasingly using structure-based approaches. Many of these structural insights for therapeutic lead development have been obtained from co-crystal structures of nAChR agonists and antagonists with the acetylcholine binding protein (AChBP). AChBP is a water soluble, structural and functional homolog of the extracellular, ligand-binding domain of nAChRs. Currently, AChBPs are recombinantly expressed in eukaryotic expression systems for structural and biophysical studies. Here, we report the establishment of an Escherichia coli (E. coli) expression system that significantly reduces the cost and time of production compared to the existing expression systems. E. coli can efficiently express unglycosylated AChBP for crystallography and makes the expression of isotopically labelled forms feasible for NMR. We used a pHUE vector containing an N-terminal His-tagged ubiquitin fusion protein to facilitate AChBP expression in the soluble fractions, and thus avoid the need to recover protein from inclusion bodies. The purified protein yield obtained from the E. coli expression system is comparable to that obtained from existing AChBP expression systems. E. coli expressed AChBP bound nAChR agonists and antagonists with affinities matching those previously reported. Thus, the E. coli expression system significantly simplifies the expression and purification of functional AChBP for structural and biophysical studies.
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: HERDC Pre-Audit
Institute for Molecular Bioscience - Publications
 
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Created: Tue, 05 Jul 2016, 13:34:17 EST by Susan Allen on behalf of Institute for Molecular Bioscience