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. 1993 Oct;65(4):1740–1748. doi: 10.1016/S0006-3495(93)81202-5

Substitution of a hydrophobic residue alters the conformational stability of Shaker K+ channels during gating and assembly.

K McCormack 1, L Lin 1, F J Sigworth 1
PMCID: PMC1225901  PMID: 8274662

Abstract

A leucine residue at position 370 (L370) in 29-4 Shaker K+ channels resides within two overlapping sequence motifs conserved among most voltage-gated channels: the S4 segment and a leucine heptad repeat. Here we investigate the effects observed upon substitution of L370 with many other uncharged amino acid residues. We find that smaller or more hydrophilic residues produce greater alterations in both activation and inactivation gating than does substitution with other large hydrophobic residues. In addition, subunits containing less conservative substitutions at position 370 are restricted in their assembly with wild-type subunits and are unlikely to form homomultimeric channel complexes. Consistent with the idea that L370 influences the tertiary structure of these channels, the results indicate that L370 undergoes specific hydrophobic interactions during the conformational transitions of gating; similar interactions may take place during the folding, insertion, or assembly of Shaker K+ channel subunits.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Abel T., Maniatis T. Gene regulation. Action of leucine zippers. Nature. 1989 Sep 7;341(6237):24–25. doi: 10.1038/341024a0. [DOI] [PubMed] [Google Scholar]
  2. Auld V. J., Goldin A. L., Krafte D. S., Catterall W. A., Lester H. A., Davidson N., Dunn R. J. A neutral amino acid change in segment IIS4 dramatically alters the gating properties of the voltage-dependent sodium channel. Proc Natl Acad Sci U S A. 1990 Jan;87(1):323–327. doi: 10.1073/pnas.87.1.323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bezanilla F., Perozo E., Papazian D. M., Stefani E. Molecular basis of gating charge immobilization in Shaker potassium channels. Science. 1991 Nov 1;254(5032):679–683. doi: 10.1126/science.1948047. [DOI] [PubMed] [Google Scholar]
  4. Bowie J. U., Reidhaar-Olson J. F., Lim W. A., Sauer R. T. Deciphering the message in protein sequences: tolerance to amino acid substitutions. Science. 1990 Mar 16;247(4948):1306–1310. doi: 10.1126/science.2315699. [DOI] [PubMed] [Google Scholar]
  5. Butler A., Wei A. G., Baker K., Salkoff L. A family of putative potassium channel genes in Drosophila. Science. 1989 Feb 17;243(4893):943–947. doi: 10.1126/science.2493160. [DOI] [PubMed] [Google Scholar]
  6. Cess R. D., Potter G. L., Blanchet J. P., Boer G. J., Ghan S. J., Kiehl J. T., LE Treut H., Li Z. X., Liang X. Z., Mitchell J. F., Morcrette J. J., Randall D. A., Riches M. R., Roeckner E., Schlese U., Slingo A., Taylor K. E., Washington W. M., Wetherald R. T., Yagai I. Interpretation of cloud-climate feedback as produced by 14 atmospheric general circulation models. Science. 1989 Aug 4;245(4917):513–516. doi: 10.1126/science.245.4917.513. [DOI] [PubMed] [Google Scholar]
  7. Engelman D. M., Steitz T. A., Goldman A. Identifying nonpolar transbilayer helices in amino acid sequences of membrane proteins. Annu Rev Biophys Biophys Chem. 1986;15:321–353. doi: 10.1146/annurev.bb.15.060186.001541. [DOI] [PubMed] [Google Scholar]
  8. Eriksson A. E., Baase W. A., Zhang X. J., Heinz D. W., Blaber M., Baldwin E. P., Matthews B. W. Response of a protein structure to cavity-creating mutations and its relation to the hydrophobic effect. Science. 1992 Jan 10;255(5041):178–183. doi: 10.1126/science.1553543. [DOI] [PubMed] [Google Scholar]
  9. Go M., Miyazawa S. Relationship between mutability, polarity and exteriority of amino acid residues in protein evolution. Int J Pept Protein Res. 1980 Mar;15(3):211–224. doi: 10.1111/j.1399-3011.1980.tb02570.x. [DOI] [PubMed] [Google Scholar]
  10. Hartmann H. A., Kirsch G. E., Drewe J. A., Taglialatela M., Joho R. H., Brown A. M. Exchange of conduction pathways between two related K+ channels. Science. 1991 Feb 22;251(4996):942–944. doi: 10.1126/science.2000495. [DOI] [PubMed] [Google Scholar]
  11. Hoshi T., Zagotta W. N., Aldrich R. W. Biophysical and molecular mechanisms of Shaker potassium channel inactivation. Science. 1990 Oct 26;250(4980):533–538. doi: 10.1126/science.2122519. [DOI] [PubMed] [Google Scholar]
  12. Isacoff E. Y., Jan Y. N., Jan L. Y. Putative receptor for the cytoplasmic inactivation gate in the Shaker K+ channel. Nature. 1991 Sep 5;353(6339):86–90. doi: 10.1038/353086a0. [DOI] [PubMed] [Google Scholar]
  13. Iverson L. E., Rudy B. The role of the divergent amino and carboxyl domains on the inactivation properties of potassium channels derived from the Shaker gene of Drosophila. J Neurosci. 1990 Sep;10(9):2903–2916. doi: 10.1523/JNEUROSCI.10-09-02903.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Landschulz W. H., Johnson P. F., McKnight S. L. The DNA binding domain of the rat liver nuclear protein C/EBP is bipartite. Science. 1989 Mar 31;243(4899):1681–1688. doi: 10.1126/science.2494700. [DOI] [PubMed] [Google Scholar]
  15. Li M., Jan Y. N., Jan L. Y. Specification of subunit assembly by the hydrophilic amino-terminal domain of the Shaker potassium channel. Science. 1992 Aug 28;257(5074):1225–1230. doi: 10.1126/science.1519059. [DOI] [PubMed] [Google Scholar]
  16. Liman E. R., Hess P., Weaver F., Koren G. Voltage-sensing residues in the S4 region of a mammalian K+ channel. Nature. 1991 Oct 24;353(6346):752–756. doi: 10.1038/353752a0. [DOI] [PubMed] [Google Scholar]
  17. Liman E. R., Tytgat J., Hess P. Subunit stoichiometry of a mammalian K+ channel determined by construction of multimeric cDNAs. Neuron. 1992 Nov;9(5):861–871. doi: 10.1016/0896-6273(92)90239-a. [DOI] [PubMed] [Google Scholar]
  18. Logothetis D. E., Movahedi S., Satler C., Lindpaintner K., Nadal-Ginard B. Incremental reductions of positive charge within the S4 region of a voltage-gated K+ channel result in corresponding decreases in gating charge. Neuron. 1992 Mar;8(3):531–540. doi: 10.1016/0896-6273(92)90281-h. [DOI] [PubMed] [Google Scholar]
  19. MacKinnon R. Determination of the subunit stoichiometry of a voltage-activated potassium channel. Nature. 1991 Mar 21;350(6315):232–235. doi: 10.1038/350232a0. [DOI] [PubMed] [Google Scholar]
  20. McCormack K., Lin J. W., Iverson L. E., Rudy B. Shaker K+ channel subunits from heteromultimeric channels with novel functional properties. Biochem Biophys Res Commun. 1990 Sep 28;171(3):1361–1371. doi: 10.1016/0006-291x(90)90836-c. [DOI] [PubMed] [Google Scholar]
  21. McCormack K., Lin L., Iverson L. E., Tanouye M. A., Sigworth F. J. Tandem linkage of Shaker K+ channel subunits does not ensure the stoichiometry of expressed channels. Biophys J. 1992 Nov;63(5):1406–1411. doi: 10.1016/S0006-3495(92)81703-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. McCormack K., Tanouye M. A., Iverson L. E., Lin J. W., Ramaswami M., McCormack T., Campanelli J. T., Mathew M. K., Rudy B. A role for hydrophobic residues in the voltage-dependent gating of Shaker K+ channels. Proc Natl Acad Sci U S A. 1991 Apr 1;88(7):2931–2935. doi: 10.1073/pnas.88.7.2931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mendel D., Ellman J. A., Chang Z., Veenstra D. L., Kollman P. A., Schultz P. G. Probing protein stability with unnatural amino acids. Science. 1992 Jun 26;256(5065):1798–1802. doi: 10.1126/science.1615324. [DOI] [PubMed] [Google Scholar]
  24. Noda M., Ikeda T., Kayano T., Suzuki H., Takeshima H., Kurasaki M., Takahashi H., Numa S. Existence of distinct sodium channel messenger RNAs in rat brain. Nature. 1986 Mar 13;320(6058):188–192. doi: 10.1038/320188a0. [DOI] [PubMed] [Google Scholar]
  25. O'Shea E. K., Klemm J. D., Kim P. S., Alber T. X-ray structure of the GCN4 leucine zipper, a two-stranded, parallel coiled coil. Science. 1991 Oct 25;254(5031):539–544. doi: 10.1126/science.1948029. [DOI] [PubMed] [Google Scholar]
  26. Papazian D. M., Timpe L. C., Jan Y. N., Jan L. Y. Alteration of voltage-dependence of Shaker potassium channel by mutations in the S4 sequence. Nature. 1991 Jan 24;349(6307):305–310. doi: 10.1038/349305a0. [DOI] [PubMed] [Google Scholar]
  27. Rees D. C., DeAntonio L., Eisenberg D. Hydrophobic organization of membrane proteins. Science. 1989 Aug 4;245(4917):510–513. doi: 10.1126/science.2667138. [DOI] [PubMed] [Google Scholar]
  28. Schoppa N. E., McCormack K., Tanouye M. A., Sigworth F. J. The size of gating charge in wild-type and mutant Shaker potassium channels. Science. 1992 Mar 27;255(5052):1712–1715. doi: 10.1126/science.1553560. [DOI] [PubMed] [Google Scholar]
  29. Stühmer W., Conti F., Stocker M., Pongs O., Heinemann S. H. Gating currents of inactivating and non-inactivating potassium channels expressed in Xenopus oocytes. Pflugers Arch. 1991 May;418(4):423–429. doi: 10.1007/BF00550881. [DOI] [PubMed] [Google Scholar]
  30. Stühmer W., Conti F., Suzuki H., Wang X. D., Noda M., Yahagi N., Kubo H., Numa S. Structural parts involved in activation and inactivation of the sodium channel. Nature. 1989 Jun 22;339(6226):597–603. doi: 10.1038/339597a0. [DOI] [PubMed] [Google Scholar]
  31. Tanabe T., Takeshima H., Mikami A., Flockerzi V., Takahashi H., Kangawa K., Kojima M., Matsuo H., Hirose T., Numa S. Primary structure of the receptor for calcium channel blockers from skeletal muscle. Nature. 1987 Jul 23;328(6128):313–318. doi: 10.1038/328313a0. [DOI] [PubMed] [Google Scholar]
  32. Tempel B. L., Papazian D. M., Schwarz T. L., Jan Y. N., Jan L. Y. Sequence of a probable potassium channel component encoded at Shaker locus of Drosophila. Science. 1987 Aug 14;237(4816):770–775. doi: 10.1126/science.2441471. [DOI] [PubMed] [Google Scholar]
  33. Tytgat J., Hess P. Evidence for cooperative interactions in potassium channel gating. Nature. 1992 Oct 1;359(6394):420–423. doi: 10.1038/359420a0. [DOI] [PubMed] [Google Scholar]
  34. Yellen G., Jurman M. E., Abramson T., MacKinnon R. Mutations affecting internal TEA blockade identify the probable pore-forming region of a K+ channel. Science. 1991 Feb 22;251(4996):939–942. doi: 10.1126/science.2000494. [DOI] [PubMed] [Google Scholar]
  35. Yool A. J., Schwarz T. L. Alteration of ionic selectivity of a K+ channel by mutation of the H5 region. Nature. 1991 Feb 21;349(6311):700–704. doi: 10.1038/349700a0. [DOI] [PubMed] [Google Scholar]

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