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. 1999 Oct;77(4):1960–1972. doi: 10.1016/S0006-3495(99)77037-2

Hydrophilicity of a single residue within MscL correlates with increased channel mechanosensitivity.

K Yoshimura 1, A Batiza 1, M Schroeder 1, P Blount 1, C Kung 1
PMCID: PMC1300477  PMID: 10512816

Abstract

Mechanosensitive channel large (MscL) encodes the large conductance mechanosensitive channel of the Escherichia coli inner membrane that protects bacteria from lysis upon osmotic shock. To elucidate the molecular mechanism of MscL gating, we have comprehensively substituted Gly(22) with all other common amino acids. Gly(22) was highlighted in random mutagenesis screens of E. coli MscL (, Proc. Nat. Acad. Sci. USA. 95:11471-11475). By analogy to the recently published MscL structure from Mycobacterium tuberculosis (, Science. 282:2220-2226), Gly(22) is buried within the constriction that closes the pore. Substituting Gly(22) with hydrophilic residues decreased the threshold pressure at which channels opened and uncovered an intermediate subconducting state. In contrast, hydrophobic substitutions increased the threshold pressure. Although hydrophobic substitutions had no effect on growth, similar to the effect of an MscL deletion, channel hyperactivity caused by hydrophilic substitutions correlated with decreased proliferation. These results suggest a model for gating in which Gly(22) moves from a hydrophobic, and through a hydrophilic, environment upon transition from the closed to open conformation.

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Selected References

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  1. Ajouz B., Berrier C., Garrigues A., Besnard M., Ghazi A. Release of thioredoxin via the mechanosensitive channel MscL during osmotic downshock of Escherichia coli cells. J Biol Chem. 1998 Oct 9;273(41):26670–26674. doi: 10.1074/jbc.273.41.26670. [DOI] [PubMed] [Google Scholar]
  2. Arkin I. T., Sukharev S. I., Blount P., Kung C., Brünger A. T. Helicity, membrane incorporation, orientation and thermal stability of the large conductance mechanosensitive ion channel from E. coli. Biochim Biophys Acta. 1998 Feb 2;1369(1):131–140. doi: 10.1016/s0005-2736(97)00219-8. [DOI] [PubMed] [Google Scholar]
  3. Bargmann C. I. Molecular mechanisms of mechanosensation? Cell. 1994 Sep 9;78(5):729–731. doi: 10.1016/s0092-8674(94)90382-4. [DOI] [PubMed] [Google Scholar]
  4. Berrier C., Besnard M., Ajouz B., Coulombe A., Ghazi A. Multiple mechanosensitive ion channels from Escherichia coli, activated at different thresholds of applied pressure. J Membr Biol. 1996 May;151(2):175–187. doi: 10.1007/s002329900068. [DOI] [PubMed] [Google Scholar]
  5. Berrier C., Coulombe A., Houssin C., Ghazi A. A patch-clamp study of ion channels of inner and outer membranes and of contact zones of E. coli, fused into giant liposomes. Pressure-activated channels are localized in the inner membrane. FEBS Lett. 1989 Dec 18;259(1):27–32. doi: 10.1016/0014-5793(89)81486-3. [DOI] [PubMed] [Google Scholar]
  6. Berrier C., Coulombe A., Szabo I., Zoratti M., Ghazi A. Gadolinium ion inhibits loss of metabolites induced by osmotic shock and large stretch-activated channels in bacteria. Eur J Biochem. 1992 Jun 1;206(2):559–565. doi: 10.1111/j.1432-1033.1992.tb16960.x. [DOI] [PubMed] [Google Scholar]
  7. Bloomfield V. Editorial. Biophys J. 1993 Jul;65(1):1–1. doi: 10.1016/S0006-3495(93)81020-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Blount P., Schroeder M. J., Kung C. Mutations in a bacterial mechanosensitive channel change the cellular response to osmotic stress. J Biol Chem. 1997 Dec 19;272(51):32150–32157. doi: 10.1074/jbc.272.51.32150. [DOI] [PubMed] [Google Scholar]
  9. Blount P., Sukharev S. I., Moe P. C., Martinac B., Kung C. Mechanosensitive channels of bacteria. Methods Enzymol. 1999;294:458–482. doi: 10.1016/s0076-6879(99)94027-2. [DOI] [PubMed] [Google Scholar]
  10. Blount P., Sukharev S. I., Moe P. C., Nagle S. K., Kung C. Towards an understanding of the structural and functional properties of MscL, a mechanosensitive channel in bacteria. Biol Cell. 1996;87(1-2):1–8. [PubMed] [Google Scholar]
  11. Blount P., Sukharev S. I., Moe P. C., Schroeder M. J., Guy H. R., Kung C. Membrane topology and multimeric structure of a mechanosensitive channel protein of Escherichia coli. EMBO J. 1996 Sep 16;15(18):4798–4805. [PMC free article] [PubMed] [Google Scholar]
  12. Blount P., Sukharev S. I., Schroeder M. J., Nagle S. K., Kung C. Single residue substitutions that change the gating properties of a mechanosensitive channel in Escherichia coli. Proc Natl Acad Sci U S A. 1996 Oct 15;93(21):11652–11657. doi: 10.1073/pnas.93.21.11652. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Chang G., Spencer R. H., Lee A. T., Barclay M. T., Rees D. C. Structure of the MscL homolog from Mycobacterium tuberculosis: a gated mechanosensitive ion channel. Science. 1998 Dec 18;282(5397):2220–2226. doi: 10.1126/science.282.5397.2220. [DOI] [PubMed] [Google Scholar]
  14. Christensen O. Mediation of cell volume regulation by Ca2+ influx through stretch-activated channels. Nature. 1987 Nov 5;330(6143):66–68. doi: 10.1038/330066a0. [DOI] [PubMed] [Google Scholar]
  15. Cruickshank C. C., Minchin R. F., Le Dain A. C., Martinac B. Estimation of the pore size of the large-conductance mechanosensitive ion channel of Escherichia coli. Biophys J. 1997 Oct;73(4):1925–1931. doi: 10.1016/S0006-3495(97)78223-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. French A. S. Mechanotransduction. Annu Rev Physiol. 1992;54:135–152. doi: 10.1146/annurev.ph.54.030192.001031. [DOI] [PubMed] [Google Scholar]
  17. Gu L., Liu W., Martinac B. Electromechanical coupling model of gating the large mechanosensitive ion channel (MscL) of Escherichia coli by mechanical force. Biophys J. 1998 Jun;74(6):2889–2902. doi: 10.1016/S0006-3495(98)77995-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hamill O. P., McBride D. W., Jr The pharmacology of mechanogated membrane ion channels. Pharmacol Rev. 1996 Jun;48(2):231–252. [PubMed] [Google Scholar]
  19. Häse C. C., Minchin R. F., Kloda A., Martinac B. Cross-linking studies and membrane localization and assembly of radiolabelled large mechanosensitive ion channel (MscL) of Escherichia coli. Biochem Biophys Res Commun. 1997 Mar 27;232(3):777–782. doi: 10.1006/bbrc.1997.6370. [DOI] [PubMed] [Google Scholar]
  20. Koch A. L., Woeste S. Elasticity of the sacculus of Escherichia coli. J Bacteriol. 1992 Jul;174(14):4811–4819. doi: 10.1128/jb.174.14.4811-4819.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  22. Lansman J. B., Hallam T. J., Rink T. J. Single stretch-activated ion channels in vascular endothelial cells as mechanotransducers? 1987 Feb 26-Mar 4Nature. 325(6107):811–813. doi: 10.1038/325811a0. [DOI] [PubMed] [Google Scholar]
  23. Levina N., Tötemeyer S., Stokes N. R., Louis P., Jones M. A., Booth I. R. Protection of Escherichia coli cells against extreme turgor by activation of MscS and MscL mechanosensitive channels: identification of genes required for MscS activity. EMBO J. 1999 Apr 1;18(7):1730–1737. doi: 10.1093/emboj/18.7.1730. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Martinac B., Buechner M., Delcour A. H., Adler J., Kung C. Pressure-sensitive ion channel in Escherichia coli. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2297–2301. doi: 10.1073/pnas.84.8.2297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Moe P. C., Blount P., Kung C. Functional and structural conservation in the mechanosensitive channel MscL implicates elements crucial for mechanosensation. Mol Microbiol. 1998 May;28(3):583–592. doi: 10.1046/j.1365-2958.1998.00821.x. [DOI] [PubMed] [Google Scholar]
  26. Nakamaru Y., Takahashi Y., Unemoto T., Nakamura T. Mechanosensitive channel functions to alleviate the cell lysis of marine bacterium, Vibrio alginolyticus, by osmotic downshock. FEBS Lett. 1999 Feb 12;444(2-3):170–172. doi: 10.1016/s0014-5793(99)00054-x. [DOI] [PubMed] [Google Scholar]
  27. Ou X., Blount P., Hoffman R. J., Kung C. One face of a transmembrane helix is crucial in mechanosensitive channel gating. Proc Natl Acad Sci U S A. 1998 Sep 15;95(19):11471–11475. doi: 10.1073/pnas.95.19.11471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sachs F., Morris C. E. Mechanosensitive ion channels in nonspecialized cells. Rev Physiol Biochem Pharmacol. 1998;132:1–77. doi: 10.1007/BFb0004985. [DOI] [PubMed] [Google Scholar]
  29. Sackin H. Mechanosensitive channels. Annu Rev Physiol. 1995;57:333–353. doi: 10.1146/annurev.ph.57.030195.002001. [DOI] [PubMed] [Google Scholar]
  30. Saint N., Lacapère J. J., Gu L. Q., Ghazi A., Martinac B., Rigaud J. L. A hexameric transmembrane pore revealed by two-dimensional crystallization of the large mechanosensitive ion channel (MscL) of Escherichia coli. J Biol Chem. 1998 Jun 12;273(24):14667–14670. doi: 10.1074/jbc.273.24.14667. [DOI] [PubMed] [Google Scholar]
  31. Sukharev S. I., Blount P., Martinac B., Blattner F. R., Kung C. A large-conductance mechanosensitive channel in E. coli encoded by mscL alone. Nature. 1994 Mar 17;368(6468):265–268. doi: 10.1038/368265a0. [DOI] [PubMed] [Google Scholar]
  32. Sukharev S. I., Blount P., Martinac B., Kung C. Mechanosensitive channels of Escherichia coli: the MscL gene, protein, and activities. Annu Rev Physiol. 1997;59:633–657. doi: 10.1146/annurev.physiol.59.1.633. [DOI] [PubMed] [Google Scholar]
  33. Sukharev S. I., Sigurdson W. J., Kung C., Sachs F. Energetic and spatial parameters for gating of the bacterial large conductance mechanosensitive channel, MscL. J Gen Physiol. 1999 Apr;113(4):525–540. doi: 10.1085/jgp.113.4.525. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Ubl J., Murer H., Kolb H. A. Ion channels activated by osmotic and mechanical stress in membranes of opossum kidney cells. J Membr Biol. 1988 Sep;104(3):223–232. doi: 10.1007/BF01872324. [DOI] [PubMed] [Google Scholar]

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