Ionisation behaviour and solution properties of the potassium-channel blocker ShK toxin

Jane E. Tudor, Michael W. Pennington, Raymond S. Norton

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The effects of pH, temperature and polypeptide concentration on the solution structure and side chain interactions of ShK toxin, a potassium-channel-blocking polypeptide from the sea anemone Stichodactyla helianthus, have been investigated by means of one-dimensional and two-dimensional 1H-NMR spectroscopy. Resonance assignments have been obtained for most protons in the molecule, and for the α and β carbon atoms. The lack of concentration dependence of the 1H chemical shifts and linewidths indicates that self-association is not significant and cannot account for the sheet-like structure near the N terminus. The structure is stable to high temperature, showing little change even at 353 K. This stability allowed backbone-amide temperature coefficients to be interpreted, and the correlation of these values with hydrogen bonds observed in the structures and with solvent exchange rates is discussed. pK(a) values have been measured for Asp5, His19 and Tyr23, and the contributions to these pK(a) values from other residues investigated using the analogues R11Q (denoting substitution of Arg11 with Gln), R11E, H19K, K22A, Y23A and K30A. These results show that Asp5 (pK(a) 2.8) makes an electrostatic interaction with Lys30, which may be partially responsible for the importance of these side chains in the folding of synthetic toxin. The phenolic pK(a) of Tyr23 is reduced to 8.7 in the native toxin, as a result of interactions with the positively charged side chains of Arg11 and to a lesser extent Lys22. Several hydrogen bonds between the Arg11 guanidino group and the Tyr23 phenolic group are found in the solution structures. As these three residues are implicated in the tight binding of ShK toxin to the T-lymphocyte voltage-gated potassium channel Kv1.3, their close interactions should be taken into account in models of binding of this toxin to the pore and vestibule of this and other potassium channels.

Original languageEnglish
Pages (from-to)133-141
Number of pages9
JournalEuropean Journal of Biochemistry
Issue number1-2
Publication statusPublished - 15 Jan 1998
Externally publishedYes


  • Electrostatic interaction
  • NMR
  • pH titration
  • Protein structure
  • ShK toxin

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