Abstract
Large-conductance (BK type) Ca(2+)-dependent K(+) channels are essential for modulating muscle contraction and neuronal activities such as synaptic transmission and hearing. BK channels are activated by membrane depolarization and intracellular Ca(2+) and Mg(2+) (refs 6-10). The energy provided by voltage, Ca(2+) and Mg(2+) binding are additive in activating the channel, suggesting that these signals open the activation gate through independent pathways. Here we report a molecular investigation of a Mg(2+)-dependent activation mechanism. Using a combined site-directed mutagenesis and structural analysis, we demonstrate that a structurally new Mg(2+)-binding site in the RCK/Rossman fold domain -- an intracellular structural motif that immediately follows the activation gate S6 helix -- is responsible for Mg(2+)-dependent activation. Mutations that impair or abolish Mg(2+) sensitivity do not affect Ca(2+) sensitivity, and vice versa. These results indicate distinct structural pathways for Mg(2+)- and Ca(2+)-dependent activation and suggest a possible mechanism for the coupling between Mg(2+) binding and channel opening.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Amino Acid Motifs
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Amino Acid Sequence
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Animals
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Binding Sites
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Calcium / metabolism
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Calcium / pharmacology
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Drosophila Proteins / chemistry
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Drosophila Proteins / genetics
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Drosophila Proteins / metabolism
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Electric Conductivity
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Escherichia coli Proteins / chemistry
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Escherichia coli Proteins / genetics
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Escherichia coli Proteins / metabolism
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Ion Channel Gating / drug effects*
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Large-Conductance Calcium-Activated Potassium Channel alpha Subunits
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Large-Conductance Calcium-Activated Potassium Channels
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Magnesium / metabolism
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Magnesium / pharmacology*
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Mice
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Models, Molecular
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Molecular Sequence Data
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Mutagenesis, Site-Directed
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Mutation
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Potassium / metabolism
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Potassium Channels / chemistry*
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Potassium Channels / genetics
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Potassium Channels / metabolism*
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Potassium Channels, Calcium-Activated / chemistry*
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Potassium Channels, Calcium-Activated / genetics
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Potassium Channels, Calcium-Activated / metabolism*
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Protein Structure, Tertiary
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
Substances
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Drosophila Proteins
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Escherichia coli Proteins
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Kcnma1 protein, mouse
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Kcnu1 protein, mouse
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Large-Conductance Calcium-Activated Potassium Channel alpha Subunits
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Large-Conductance Calcium-Activated Potassium Channels
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Potassium Channels
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Potassium Channels, Calcium-Activated
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Recombinant Fusion Proteins
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Magnesium
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Potassium
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Calcium