Solution structure of the potassium channel inhibitor agitoxin 2: Caliper for probing channel geometry

Authors

  • Andrzej M. Krezel,

    1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115
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    • A.M. Krezel and C. Kasibhatla contributed equally to this work.

  • Chandrasekhar Kasibhatla,

    1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115
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    • A.M. Krezel and C. Kasibhatla contributed equally to this work.

  • Patricia Hidalgo,

    1. Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115
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  • Roderick Mackinnon,

    1. Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115
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  • Gerhard Wagner

    Corresponding author
    1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115
    • Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115
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Abstract

The structure of the potassium channel blocker agitoxin 2 was solved by solution NMR methods. The structure consists of a triple-stranded antiparallel β-sheet and a single helix covering one face of the β-sheet. The cysteine side chains connecting the β-sheet and the helix form the core of the molecule. One edge of the β-sheet and the adjacent face of the helix form the interface with the Shaker K+ channel. The fold of agitoxin is homologous to the previously determined folds of scorpion venom toxins. However, agitoxin 2 differs significantly from the other channel blockers in the specificity of its interactions. This study was thus focused on a precise characterization of the surface residues at the face of the protein interacting with the Shaker K+ channel. The rigid toxin molecule can be used to estimate dimensions of the potassium channel. Surface-exposed residues, Arg24, Lys27, and Arg31 of the β-sheet, have been identified from mutagenesis studies as functionally important for blocking the Shaker K+ channel. The sequential and spatial locations of Arg24 and Arg31 are not conserved among the homologous toxins. Knowledge on the details of the channel-binding sites of agitoxin 2 formed a basis for site-directed mutagenesis studies of the toxin and the K+ channel sequences. Observed interactions between mutated toxin and channel are being used to elucidate the channel structure and mechanisms of channel-toxin interactions.

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