Single molecule FRET reveals pore size and opening mechanism of a mechano-sensitive ion channel

Abstract

The mechanosensitive channel of large conductance, which serves as a model system for mechanosensitive channels, has previously been crystallized in the closed form, but not in the open form. Ensemble measurements and electrophysiological sieving experiments show that the open-diameter of the channel pore is >25 Å, but the exact size and whether the conformational change follows a helix-tilt or barrel-stave model are unclear. Here we report measurements of the distance changes on liposome-reconstituted MscL transmembrane α-helices, using a ‘virtual sorting’ single-molecule fluorescence energy transfer. We observed directly that the channel opens via the helix-tilt model and the open pore reaches 2.8 nm in diameter. In addition, based on the measurements, we developed a molecular dynamics model of the channel structure in the open state which confirms our direct observations.

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References 65

1. 1.

   Akaike H 1974 A new look at the statistical model identification. IEEE Transactions on Automatic Control19:716–723.

   Not in our library10.1109/tac.1974.1100705

2. 2.

   Akyuz N Altman RB Blanchard SC Boudker O 2013 Transport dynamics in a glutamate transporter homologue. Nature502:114–118.

   Not in our library10.1038/nature12265

3. 3.

   Andrews DL Demidov AA 1999 Resonance energy transfer. 1st edition Guildford and King’s Lynn: Wiley

   Not in our library

4. 4.

   Anishkin A Gendel V Sharifi NA Chiang C-S Shirinian L Guy HR Sukharev S 2003 On the conformation of the COOH-terminal domain of the large mechanosensitive channel MscL. The Journal of General Physiology121:227–244.

   Not in our library10.1085/jgp.20028768

5. 5.

   Árnadóttir J Chalfie M 2010 Eukaryotic mechanosensitive channels. Annual Review of Biophysics39:111–137.

   Not in our library10.1146/annurev.biophys.37.032807.125836

6. 6.

   Best RB Zhu X Shim J Lopes PEM Mittal J Feig M MacKerell AD 2012 Optimization of the additive CHARMM all-atom protein force field targeting improved sampling of the backbone ϕ, ψ and side-chain χ1 and χ2 dihedral angles. Journal of Chemical Theory and Computation8:3257–3273.

   Not in our library10.1021/ct300400x

7. 7.

   Betanzos M Chiang C-S Guy HR Sukharev S 2002 A large iris-like expansion of a mechanosensitive channel protein induced by membrane tension. Nature Structural & Molecular Biology9:704–710.

   Not in our library10.1038/nsb828

8. 8.

   Booth IR Blount P 2012 The MscS and MscL families of mechanosensitive channels act as microbial emergency release valves. Journal of Bacteriology194:4802–4809.

   Not in our library10.1128/jb.00576-12

9. 9.

   Brünger AT Clore GM Gronenborn AM Karplus M 1986 Three-dimensional structure of proteins determined by molecular dynamics with interproton distance restraints: application to crambin. Proceedings of the National Academy of Sciences of the United States of America83:3801–3805.

   Not in our library10.1073/pnas.83.11.3801

10. 10.

    Cha A Snyder GE Selvin PR Bezanilla F 1999 Atomic scale movement of the voltage-sensing region in a potassium channel measured via spectroscopy. Nature402:809–813.

    Not in our library10.1038/45552

11. 11.

    Chalfie M 2009 Neurosensory mechanotransduction. Nature Reviews Molecular Cell Biology10:44–52.

    Not in our library10.1038/nrm2595

12. 12.

    Chan K-Y Gumbart J McGreevy R Watermeyer JM Sewell BT Schulten K 2011 Symmetry-restrained flexible fitting for symmetric EM Maps. Structure19:1211–1218.

    Not in our library10.1016/j.str.2011.07.017

13. 13.

    Chang G Spencer RH Lee AT Barclay MT Rees DC 1998 Structure of the MscL Homolog from Mycobacterium tuberculosis: a gated mechanosensitive ion channel. Science282:2220–2226.

    Not in our library10.1126/science.282.5397.2220

14. 14.

    Choi UB Strop P Vrljic M Chu S Brunger AT Weninger KR 2010 Single-molecule FRET-derived model of the synaptotagmin 1-SNARE fusion complex. Nature Structural & Molecular Biology17:318–324.

    Not in our library10.1038/nsmb.1763

15. 15.

    Chung M Lowe RD Chan Y-HM Ganesan PV Boxer SG 2009 DNA-tethered membranes formed by giant vesicle rupture. Journal of Structural Biology168:190–199.

    Not in our library10.1016/j.jsb.2009.06.015

16. 16.

    Clegg RM 1992 Fluorescence resonance energy transfer and nucleic acids. In: David MJ Lilley JED, editors. Methods in Enzymology. Academic Press p. 353–388.

    Not in our library10.1016/0076-6879(92)11020-j

17. 17.

    Corry B Hurst AC Pal P Nomura T Rigby P Martinac B 2010 An improved open-channel structure of MscL determined from FRET confocal microscopy and simulation. The Journal of General Physiology136:483–494.

    Not in our library10.1085/jgp.200910376

18. 18.

    Corry B Jayatilaka D Rigby P 2005a A flexible approach to the calculation of resonance energy transfer efficiency between multiple donors and acceptors in complexgeometries. Biophysical Journal89:3822–3836.

    Not in our library10.1529/biophysj.105.069351

19. 19.

    Corry B Rigby P Liu Z-W Martinac B 2005b Conformational changes involved in MscL channel gating measured using FRET spectroscopy. Biophysical Journal89:L49–L51.

    Not in our library10.1529/biophysj.105.072009

20. 20.

    Cruickshank CC Minchin RF Le Dain AC Martinac B 1997 Estimation of the pore size of the large-conductance mechanosensitive ion channel of Escherichia coli. Biophysical Journal73:1925–1931.

    Not in our library10.1016/s0006-3495(97)78223-7

21. 21.

    Dale RE Eisinger J Blumberg WE 1979 The orientational freedom of molecular probes. The orientation factor in intramolecular energy transfer. Biophysical Journal26:161–193.

    Not in our library10.1016/s0006-3495(79)85243-1

22. 22.

    Deplazes E Louhivuori M Jayatilaka D Marrink SJ Corry B 2012 Structural investigation of MscL gating using experimental data and coarse grained MD simulations. PLOS Computational Biology8:e1002683.

    Not in our library10.1371/journal.pcbi.1002683

23. 23.

    Fery-Forgues S Lavabre D 1999 Are fluorescence quantum yields so tricky to measure? A demonstration using familiar stationery products. Journal of Chemical Education76:1260.

    Not in our library10.1021/ed076p1260

24. 24.

    Förster T 1948 Zwischenmolekulare energiewanderung und fluoreszenz. Annalen der Physik437:55–75.

    Not in our library10.1002/andp.19484370105

25. 25.

    Gandhi CS Walton TA Rees DC 2011 OCAM: a new tool for studying the oligomeric diversity of MscL channels. Protein Science20:313–326.

    Not in our library10.1002/pro.562

26. 26.

    Granier S Kim S Shafer AM Ratnala VRP Fung JJ Zare RN Kobilka B 2007 Structure and conformational changes in the C-terminal domain of the β2-adrenoceptor. Journal of Biological Chemistry282:13895–13905.

    Not in our library10.1074/jbc.m611904200

27. 27.

    Hamill OP Martinac B 2001 Molecular basis of mechanotransduction in living cells. Physiological Reviews81:685–740

    Not in our library10.1152/physrev.2001.81.2.685

28. 28.
    Avogadro: an advanced semantic chemical editor, visualization, and analysis platform · 2012

    No notice
29. 29.

    Haswell ES Phillips R Rees DC 2011 Mechanosensitive channels: what can they do and how do they do it?Structure19:1356–1369.

    Not in our library10.1016/j.str.2011.09.005

30. 30.

    Humphrey W Dalke A Schulten K 1996 VMD: visual molecular dynamics. Journal of Molecular Graphics14:33–38, 27–28.

    Not in our library10.1016/0263-7855(96)00018-5

31. 31.

    Jae M 2003 Tutorial on maximum likelihood estimation. Journal of Mathematical Psychology47:90–100.

    Not in our library10.1016/s0022-2496(02)00028-7

32. 32.

    Iqbal A Arslan S Okumus B Wilson TJ Giraud G Norman DG Ha T Lilley DMJ 2008 Orientation dependence in fluorescent energy transfer between Cy3 and Cy5 terminally attached to double-stranded nucleic acids. Proceedings of the Montana Academy of Sciences105:11176–11181.

    Not in our library10.1073/pnas.0801707105

33. 33.

    Jäger M Nir E Weiss S 2006 Site-specific labeling of proteins for single-molecule FRET by combining chemical and enzymatic modification. Protein Science15:640–646.

    Not in our library10.1110/ps.051851506

34. 34.

    Jorgensen WL Chandrasekhar J Madura JD Impey RW Klein ML 1983 Comparison of simple potential functions for simulating liquid water. The Journal of Chemical Physics79:926–935.

    Not in our library10.1063/1.445869

35. 35.

    Kim Y Ho SO Gassman NR Korlann Y Landorf EV Collart FR Weiss S 2008 Efficient site-specific labeling of proteins via cysteines. Bioconjugate Chemistry19:786–791.

    Not in our library10.1021/bc7002499

36. 36.

    Lakowicz JR 1999 Principles of fluorescence spectroscopy. 2nd edition Springer

    Not in our library

37. 37.

    Liu Z Gandhi CS Rees DC 2009 Structure of a tetrameric MscL in an expanded intermediate state. Nature461:120–124.

    Not in our library10.1038/nature08277

38. 38.

    Louhivuori M Risselada HJ van der Giessen E Marrink SJ 2010 Release of content through mechano-sensitive gates in pressurized liposomes. Proceedings of the National Academy of Sciences of the United States of America107:19856–19860.

    Not in our library10.1073/pnas.1001316107

39. 39.

    Majumdar DS Smirnova I Kasho V Nir E Kong X Weiss S Kaback HR 2007 Single-molecule FRET reveals sugar-induced conformational dynamics in LacY. Proceedings of the National Academy of Sciences of the United States of America104:12640–12645.

    Not in our library10.1073/pnas.0700969104

40. 40.

    Marras SAE Kramer FR Tyagi S 2002 Efficiencies of fluorescence resonance energy transfer and contact–mediated quenching in oligonucleotide probes. Nucleic Acids Research30:e122.

    Not in our library10.1093/nar/gnf121

Cited by papers we hold 3

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