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J Gen Physiol. 2015 Oct;146(4):323-40. doi: 10.1085/jgp.201511478.

A chimeric prokaryotic pentameric ligand-gated channel reveals distinct pathways of activation.

Author information

1
Department of Neuroscience, Department of Physiology and Biophysics, Department of Biochemistry, and Center for Proteomics and Bioinformatics, School of Medicine, Case Western Reserve University, Cleveland, OH 44106.
2
Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37232.
3
Department of Neuroscience, Department of Physiology and Biophysics, Department of Biochemistry, and Center for Proteomics and Bioinformatics, School of Medicine, Case Western Reserve University, Cleveland, OH 44106 Department of Neuroscience, Department of Physiology and Biophysics, Department of Biochemistry, and Center for Proteomics and Bioinformatics, School of Medicine, Case Western Reserve University, Cleveland, OH 44106 Sudha.chakrapani@case.edu.

Abstract

Recent high resolution structures of several pentameric ligand-gated ion channels have provided unprecedented details of their molecular architecture. However, the conformational dynamics and structural rearrangements that underlie gating and allosteric modulation remain poorly understood. We used a combination of electrophysiology, double electron-electron resonance (DEER) spectroscopy, and x-ray crystallography to investigate activation mechanisms in a novel functional chimera with the extracellular domain (ECD) of amine-gated Erwinia chrysanthemi ligand-gated ion channel, which is activated by primary amines, and the transmembrane domain of Gloeobacter violaceus ligand-gated ion channel, which is activated by protons. We found that the chimera was independently gated by primary amines and by protons. The crystal structure of the chimera in its resting state, at pH 7.0 and in the absence of primary amines, revealed a closed-pore conformation and an ECD that is twisted with respect to the transmembrane region. Amine- and pH-induced conformational changes measured by DEER spectroscopy showed that the chimera exhibits a dual mode of gating that preserves the distinct conformational changes of the parent channels. Collectively, our findings shed light on both conserved and divergent features of gating mechanisms in this class of channels, and will facilitate the design of better allosteric modulators.

PMID:
26415570
PMCID:
PMC4586589
DOI:
10.1085/jgp.201511478
[Indexed for MEDLINE]
Free PMC Article

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