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Engineering a surrogate human heteromeric α/β glycine receptor orthosteric site exploiting the structural homology and stability of acetylcholine-binding protein

DOI: 10.1107/S205225251901114X DOI Help

Authors: Alice Dawson (University of Dundee) , Paul Trumper (University of Dundee) , Juliana Oliveira De Souza (University of Dundee) , Holly Parker (University of Dundee) , Mathew J. Jones (University of Dundee) , Tim G. Hales (University of Dundee) , William N. Hunter (University of Dundee)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Iucrj , VOL 6

State: Published (Approved)
Published: November 2019
Diamond Proposal Number(s): 19844

Open Access Open Access

Abstract: Protein-engineering methods have been exploited to produce a surrogate system for the extracellular neurotransmitter-binding site of a heteromeric human ligand-gated ion channel, the glycine receptor. This approach circumvents two major issues: the inherent experimental difficulties in working with a membrane-bound ion channel and the complication that a heteromeric assembly is necessary to create a key, physiologically relevant binding site. Residues that form the orthosteric site in a highly stable ortholog, acetylcholine-binding protein, were selected for substitution. Recombinant proteins were prepared and characterized in stepwise fashion exploiting a range of biophysical techniques, including X-ray crystallography, married to the use of selected chemical probes. The decision making and development of the surrogate, which is termed a glycine-binding protein, are described, and comparisons are provided with wild-type and homomeric systems that establish features of molecular recognition in the binding site and the confidence that the system is suited for use in early-stage drug discovery targeting a heteromeric α/β glycine receptor.

Journal Keywords: acetylcholine-binding protein; crystal structures; glycine receptor; ligand-gated ion channel; nicotine; strychnine; tropisetron

Subject Areas: Biology and Bio-materials

Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)

Other Facilities: ESRF ID23

Added On: 29/09/2019 22:17


Discipline Tags:

Health & Wellbeing Neurology Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)