Abstract
G protein-coupled potassium channels (GIRK/Kir3.x) are key determinants that translate inhibitory chemical neurotransmission into changes in cellular excitability. To understand the mechanism of channel activation by G proteins, it is necessary to define the structural rearrangements in the channel that result from interaction with Gbetagamma subunits. In this study we used a combination of fluorescence spectroscopy and through-the-objective total internal reflection microscopy to monitor the conformational rearrangements associated with the activation of GIRK channels in single intact cells. We detect activation-induced changes in FRET consistent with a rotation and expansion of the termini along the central axis of the channel. We propose that this rotation and expansion of the termini drives the channel to open by bending and possibly rotating the second transmembrane segment.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Bacterial Proteins / genetics
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Cell Membrane / metabolism
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Cell Membrane / ultrastructure
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Fluorescence Polarization
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Fluorescence Resonance Energy Transfer*
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Fluorescent Dyes
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G Protein-Coupled Inwardly-Rectifying Potassium Channels
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Green Fluorescent Proteins
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Ion Channel Gating / physiology*
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Luminescent Proteins / genetics
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Microscopy, Fluorescence / instrumentation
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Microscopy, Fluorescence / methods
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Models, Molecular
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Oocytes / metabolism
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Patch-Clamp Techniques
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Potassium Channels / genetics
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Potassium Channels / metabolism*
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Potassium Channels, Inwardly Rectifying*
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Protein Conformation
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Protein Subunits / genetics
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Protein Subunits / metabolism
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Signal Transduction / physiology
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Transfection
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Xenopus laevis
Substances
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Bacterial Proteins
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Fluorescent Dyes
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G Protein-Coupled Inwardly-Rectifying Potassium Channels
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Luminescent Proteins
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Potassium Channels
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Potassium Channels, Inwardly Rectifying
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Protein Subunits
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Recombinant Fusion Proteins
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yellow fluorescent protein, Bacteria
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Green Fluorescent Proteins