Skip to main content
SpringerLink
Log in

Sizing Channels with Neutral Polymers

  • Conference paper
Structure and Dynamics of Confined Polymers

Part of the book series: NATO Science Series ((ASHT,volume 87))

Abstract

We have developed a method for determining the physical dimensions of nanometer scale pores formed by protein ion channels. This was possible because of the availability of a wide range of size-selected nonelectrolyte polymers of poly(ethylene glycol), PEG, and because PEG decreases the bulk conductivity of ionic solutions. The method is simple. PEGs that are sufficiently small enter the channel’s pore and decrease the channel’s ionic conductance. PEGs that are larger than the pore’s diameter rarely partition into the pore and therefore do not decrease the channel conductance. Thus, the dependence of the channel conductance on the PEG molecular weight determines the pore’s PEG molecular weight cut-off, and by inference, the pore’s radius. We recently extended the technique to determine the shape of a channel’s lumen including the sizes of both openings and the size and location of constrictions inside the pore. We discuss here the details of the method, the properties of PEG, and some limitations of using the technique to determine channel size.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 169.00
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 219.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 219.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. North C.L., Barranger-Mathys, M. and Cafiso, D. (1995) Membrane orientation of the N-terminal segment of alamethicin determined by solid-State 15N NMR. Biophys. J. 69, 2392–2397.

    Article  ADS  Google Scholar 

  2. Opella S.J. (1994). Solid-state NMR structural studies of proteins. Ann. Rev. Phys. Chem. 45, 659–683.

    Article  ADS  Google Scholar 

  3. Altenbach, C., Froncisz, W., Hubbell, W. and Hyde, J. (1988) The orientation of membrane-bound, spin-labeled mellitin as determined by electron-paramagnetic resaturation recovery measurements. Biophys. J. 53, A94–A94.

    Google Scholar 

  4. Cafiso, D.S. (1994) Alamethicin - a peptide model for voltage gating and protein membrane interactions. Ann. Rev. Biophys. Biomol. Struct. 23,141–165.

    Article  Google Scholar 

  5. Barranger-Mathys, M. and Cafiso, D.S. (1996) Membrane Structure of Voltage-Gated Channel Forming Peptides by Site-Directed Spin-Labeling. Biochemistry 35, 498–505.

    Article  Google Scholar 

  6. Tsukihara, T. and Aoyama, H. (2000) Membrane protein assemblies - towards atomic resolution analysis. Curr. Opin. Struct. Biol. 10, 208–212.

    Article  Google Scholar 

  7. Fleming, K.G. (2000) Riding the wave: structural and energetic principles of helical membrane proteins.Curr. Opin. Biotechnol.11,67–71.

    Article  Google Scholar 

  8. Sakai, H. and Tsukihara, T. (1998). Structures of membrane proteins determined at atomic resolution.J Biochem. (Tokyo) 124,1051–1059.

    Article  Google Scholar 

  9. Ostermeier, C. and Michel, H. (1997) Crystallization of membrane proteins.Cure Opin. Struct. Biol.7, 697–701; Beauchamp, J.C. and Isaacs, N.W. (1999) Methods for X-ray diffraction analysis of macromolecular structures.Curr. Opin. Chem. Biol. 3, 525–529.

    Google Scholar 

  10. Akabas, M.H., Stauffer, D.A., Xu, M. and Karlin, A. (1992) Acetylcholine-receptor channel structure probed in cysteine-substitution mutantsScience 258,307–310.

    Article  ADS  Google Scholar 

  11. Zimmerberg, J. and Parsegian, V.A.P. (1986) Polymer inaccessible volume changes during opening and closing of a voltage-dependent ionic channel.Nature (London) 323, 36–39.

    Article  ADS  Google Scholar 

  12. Krasilnikov, O.V., Sabirov, R.Z., Ternovsky, V.I., Merzliak, P.G. and Muratkhodjaev, J.N. (1992a) A simple method for the determination of the pore radius of ion channels in planar lipid bilayer membranes.FEMS Microbiol. Immunol.105, 93–100.

    Article  Google Scholar 

  13. Sabirov, R.Z., Krasilnikov, O.V., Ternovsky, V.I., Merzliak P.G. and Muratkhodjaev, J.N. (1991) Influence of some nonelectrolytes on conductivity of bulk solution and conductance of ion channels. Determination of pore radius from electric measurements.Biologicheskie Membrany 8, 280–291.

    Google Scholar 

  14. Bezrukov, S.M. and Vodyanoy, I. (1993) Probing alamethicin channels with water-soluble polymers. Effect on conductance of channel states. Biophys. J 64, 16–25.

    Article  Google Scholar 

  15. Sabirov R.Z., Krasilnikov O.V., Ternovsky V.I., Merzliak P.G. (1993): Relation between ionic channel conductance and conductivity of media containing different non-electrolytes. A novel method of pore size determination.Gen. Physiol. Biophys.12, 95–111.

    Google Scholar 

  16. Hager, S.L. and MacRury, T.B. (1980) Investigation of phase-behavior and water binding in poly(alkilene oxide) solutions.J.Appl. Polym. Sci.25, 1559–1571.

    Article  Google Scholar 

  17. Tilcock, C.P.S. and Fisher D. (1982) The interaction of phospholipid membranes with poly(ethylene glycol). Vesicle aggregation and lipid exchange.Biochim. Biophys. Acta 688, 645–652.

    Article  Google Scholar 

  18. Breen J., Huis D., Bleijser J. and Leyte J.C. (1988) Solvent dynamics in aqueous PEO-salt solutions studied by nuclear magnetic relaxation.J.Chem.Soc., Faraday Trans.I 84, 293–307

    Article  Google Scholar 

  19. Antonsen, K.P., Hoffman A.S. (1992) Water structure of PEG solutions by differential scanning calorimetry measurements. In:Poly(Ethylene Glycol) Chemistry: Biotechnical and Biomedical Applications, Edited by Harris, J.M., Plenum Press, New York-London, 15–28.

    Google Scholar 

  20. Bailey F.E. and Koleske, I.V., (1976)Poly(Ethylene Oxide)Academic Press, New York

    Google Scholar 

  21. Bailey F.E. and Koleske I.V. (1991)Alkylene Oxides and Their Polymers, Marcel Dekker, New York

    Google Scholar 

  22. Harris J.M. (1992) Introduction to biotechnical and biomedical applications of poly(ethylene glycol). In: Poly(Ethylene Glycol) Chemistry: Biotechnical and Biomedical Applications, Edited by Harris, J.M., Plenum Press, New York-London, 1–13.

    Google Scholar 

  23. Harris J.M., Hundley N.H., Shannon, T.G., Struck, E.C. (1982) Poly(ethylene glycols) as soluble, recoverable, phase-transfercatalysts. J. Org. Chem. 47, 4789–4791.

    Article  Google Scholar 

  24. Korchev, Y.E., Bashford, C.L., Alder, C.M., Kasianowicz J.J. and Pasternak C.A. (1995) Low conductance states of a single ion channel are not “closed”. JMembr. Biol. 147, 233–239.

    Google Scholar 

  25. Krasilnikov, O.V., Yuldasheva L.N., Nogueira, R.A. and Rodrigues, C.G. (1995) The diameter of water pores formed by colicin la in planar lipid bilayers.Brazilian J. of Med. and Biol. Res.28, 693–698.

    Google Scholar 

  26. Bezrukov, S.M., Vodyanoy, I., Brutyan, R.A. and Kasianowicz, J.J. (1996) Dynamics and free energy of polymers partitioning into a nanoscale pore.Macromolecules 29, 8517–8522.

    Article  ADS  Google Scholar 

  27. Bezrukov, S.M. and Kasianowicz, J.J. (1997) The charge state of an ion channel controls neutral polymer entry into its pore. Eur. Biophys. J. 26, 471–476.

    Article  Google Scholar 

  28. Carneiro, C.M.M., Krasilnikov, O.V., Yuldasheva, L.N., Campos de Carvalho, A.C. and Nogueira R.A. (1997). Is the mammalian porin channel, VDAC, a perfect cylinder in high conductance state?FEBS Lett.416, 187–189.

    Article  Google Scholar 

  29. Krasilnikov, O.V., Merzlyak, P.G., Yuldasheva, L.N., Azimova, R.K. and Nogueira, R.A. (1997) Pore-forming properties of proteolytically nicked staphylococcal β-toxin. The ion channel in planar lipid bilayer membranes.Med. Microbiol. and Immunol.186, 53–61.

    Article  Google Scholar 

  30. Lee, J.C. and Lee, L.L. (1981) Preferential solvent interactions between proteins and polyethylene glycols. J. Biol. Chem. 256, 625–631.

    Google Scholar 

  31. Merzlyak, P.G., Yuldasheva, L.N., Rodrigues, C.G., Cameiro, C.M., Krasilnikov O.V. and Bezrukov, S.M. (1999) Polymeric Nonelectrolytes to probe pore geometry: Application to the β-toxin transmembrane channel.Biophys. J.77, 3023–3033.

    Article  Google Scholar 

  32. Bhakdi, S. and Tranum-Jensen, J.(1991) S. aureus β-toxin. Microbiol. Rev. 55, 733–751.

    Google Scholar 

  33. Gray, G.S. and Kehoe, M. (1984) Primary sequence of the a-toxin gene from Staphylococcus aureus Wood 46.Infect. Immunity 46, 615–618.

    Google Scholar 

  34. ] Valeva, A., Weisser, A., Walker, B., Kehoe, M., Bayley, H., Bhakdi S. and Palmer, M. (1996) Molecular architecture of a toxin pore: a 15-residue sequence lines the trans-membrane channel of Staphylococcal alpha-toxin.EMBO J.15, 1857–1864.

    Google Scholar 

  35. Vecsey-Semjen, B., Lesieur, C., Mollby, R. and vander Goot, F.G. (1997) Conformational changes due to membrane binding and channel formation by staphylococcal alpha-toxin.J. Biol. Chem.272, 5709–5717.

    Article  Google Scholar 

  36. Tomita, T., Watanabe, M. and Yasuda, T. (1992) Influence of membrane fluidity on assembly of Staphylococcus aureus a-toxin, a channel-forming protein, in liposome membrane.J Biol. Chem. 267, 13391–13397.

    Google Scholar 

  37. Krasilnikov, O.V., Temovsky, V.I., Musaev, Yu.M. and Tashmukhamedov, B.A. (1980). Influence of staphylotoxin on conductance of bilayer phospholipid membranes.Doklady AN UzSSR N7, 66–68.

    Google Scholar 

  38. [] Krasilnikov, O.V., Temovsky, V.I. and Tashmukhamedov, B.A. (1981). Properties of ion channels induced by alpha-staphylotoxin in bilayer lipid membranes.Biofisica 26, 271–275.

    Google Scholar 

  39. Gouaux, J.E., Braha, O., Hobaugh, M.R., Song, L., Cheley, S., Shustak, C. and Bayley, H. (1994). Subunit stoichiometry of staphylococcal alpha-hemolysin in crystals and on membranes: A heptameric transmembrane pore.Proc. Natl. Acad. Sci. (USA) 91, 12828–12831.

    Article  ADS  Google Scholar 

  40. Krasilnikov, O.V., Cruz, J.B.Da., Yuldasheva, L.N. and Nogueira, R.A. (1998) A novel approach to study the geometry of the water lumen ion channel. Colicin la channels in lipid bilayers.J. Membr. Biol. 161, 83–92.

    Article  Google Scholar 

  41. Krasilnikov, O.V., Muratkhodjaev, D.N. and Zitzer, A.O. (1992b) The mode of action of V.cholerae cytolysin. The influences of both erythrocytes and planar lipid bilayers.Biochim. Biophys. Acta 1111, 7–16.

    Article  Google Scholar 

  42. Song, L., Hobaugh, M.R., Shustak, C., Cheley S., Bayley, H., and Gouaux, J.E. (1996) Structure of staphylococcal ce-hemolysin, a heptameric transmembrane pore.Science 274, 1859–1866.

    Article  ADS  Google Scholar 

  43. Kasianowicz, J.J., Burden, D.L., Han, L., Cheley, S. and Bayley H. (1999) Genetically engineered metal ion binding sites on the outside of a channel’s transmembrane b-barrel.Biophys. J.76, 837–845.

    Article  Google Scholar 

  44. Arakawa, T. and Timasheff, S.N. (1985) Mechanism of poly(ethylene glycol) interaction with proteins.Biochemistry 24, 6756–6762.

    Article  Google Scholar 

  45. Lee, L.L. and Lee, J.C. (1987) Thermal stability of proteins in the presence of poly(ethylene glycols).Biochemistry 26, 7813–7819.

    Article  Google Scholar 

  46. Hammes, G.G. and Schimmel, P.R. (1967) An investigation of water-urea and waterurea-polyethylene glycol interactions.J. Am. Chem. Soc.89, 442–446.

    Article  Google Scholar 

  47. Ingham, K.C. (1977) Polyethylene glycol in aqueous solution: solvent perturbation and gel filtration studies.Arch. Biochem. Biophys.184, 59–68.

    Article  Google Scholar 

  48. Parsegian, V.A., Rand, R.P., Fuller, N.L. and Rau, D.C. (1986) Osmotic stress for the direct measurement of intermolecular forces.Method Enzymol.127, 400–416.

    Article  Google Scholar 

  49. Pohl, P., Saparov, S.M. and Antonenko, Y.N. (1997) The effect of a transmembrane osmotic flux on the ion concentration distribution in the immediate membrane vicinity measured by microelectrodes.Biophys. J.72, 1711–1718.

    Article  Google Scholar 

  50. Pohl, P., Saparov, S.M. and Antonenko, Y.N. (1998) The size of the unstirred layer as a function of the solute diffusion coefficient.Biophys. J.75, 1403–1409.

    Article  Google Scholar 

  51. Scherrer, R. and Gerhard, P. (1971) Molecular sieving by the Bacillus megaterium cell wall and protoplast.J.Bacteriol.107, 718–735.

    Google Scholar 

  52. Berestovsky, G.N., Ternovsky, V.I. and Cataev, A.A.Private communication.

    Google Scholar 

  53. Sabirov, R.Z., Krasilnikov, O.V., Temovsky, V.I., Merzliak, P.G. and Muratkhodjaev, J.N. (1991) Influence of some nonelectrolytes on conductivity of bulk solution and conductance of ion channels. Determination of pore radius from electric measurements.Biologicheskie Membrany 8, 280–291.

    Google Scholar 

  54. Sabirov, R.Z., Krasilnikov, O.V., Temovsky, V.I. and Merzliak, P.G. (1993) Relation between ionic channel conductance and conductivity of media containing different non-electrolytes. The novel method of pore size determination.Gen. Physiol. Biophys.12, 95–111.

    Google Scholar 

  55. Krasilnikov, O.V. and Sabirov, R.Z. (1992) Comparative analysis of latrotoxin channels of different conductance in planar lipid bilayers. Evidence for cluster organization.Biochim. Biophys. Acta 1112, 124–128.

    Article  Google Scholar 

  56. Temovsky, V.I. and Berestovsky, G.N. (1998) Effective diameter and structural organization of reconstituted calcium channels from the Characeae algae Nitellopsis.Membr. Cell Biol.12, 79–88.

    Google Scholar 

  57. Krasilnikov, O.V., Sabirov, R.Z., Temovsky, V.I., Merzliak, P.G. and Tashmukhamedov, B.A. (1988) Structure of ion channels induced by a-toxin from Staphylococcus aureus. Gen. Physiol. Biophys.7, 467–473.

    Google Scholar 

  58. Krasilnikov O.V., Muratkhodjaev D.N., Voronov S.E., Ezepchuk Yu.V. (1991) The ionic channels formed by cholera toxin in planar bilayer lipid membranes are entirely attributable to its B-subunit.Biochim. Biophys. Acta 1067,166–170.

    Article  Google Scholar 

  59. Zitzer, A.O., Nakisbekov, N.O., Li, A.V., Semiotrochev, V.L., Kiseliov, Yu.L., Muratkhodjaev, J.N., Krasilnikov, O.V. and Ezepchuk, Yu.V. (1993) Entero-cytolisin (EC) from Vebrio cholerae non-01 (some properties and pore-forming activity).Int. J. Med. Microbiol. Virol. Parasitol. Infect. Dis.279, 494–504.

    Google Scholar 

  60. Krasilnikov, O.V., Temovsky, V.I., Navasardyants, D.G. and Kalmykova, L.I. (1994) The characterization of ion channels formed by Pasteurella multocida dermonecrotic toxin.Med. Microbiol. and Immunol.183, 229–237.

    Article  Google Scholar 

  61. Desai S.A., Rosenberg R.L, (1997) Pore size of the malaria parasite’s nutrient channel.Proc. Natl. Acad. Sci. (USA) 94, 2045–2049.

    Article  ADS  Google Scholar 

  62. Kaulin Y.A., Schagina L.V., Bezrukov S.M., Maley V.V., Feigin A.M., Takemoto J.Y., Teeter J.H.., Brand J.G. (1998) Cluster organization of ion channels formed by the antibiotic Syringomycin E in bilayer lipid membranes.Biophys. J.74, 2918–2925.

    Article  ADS  Google Scholar 

  63. Sabirov R.Z., Tadjibaeva G. Sh., Krasilnikov O.V., El Sufi S.A.F., Tashmukhamedov B.A. (1992) Influence of the hydrophilic nonelectrolytes on the single amphotericin channels.DAN UzSSR N1, 52–54.

    Google Scholar 

  64. Temovsky V.I., Grigoriev P.A., Berestovsky G.N., Schlegel R., Dornberger K., Grafe U. (1997) Effective diameters of ion channels formed by homologs of the antibiotic chrysospermin.Membr. Cell Biol.11 497–505.

    Google Scholar 

  65. Goudet C, Benitah J.P., Milat M.L., Sentenac H., and Thibaud J.B. (1999) Cluster organization and pore structure of ion channels formed by beticolin 3, a nonpeptidic fungal toxin.Biophys. J.77, 3052–3059.

    Article  Google Scholar 

  66. Coates G.M.P., Bashford C.L., Smart O.S. (1998) Using HOLE to predict the effects of PEG’s on the conductance of a-toxin. Biochem.Soc. Trans. 26, S193.

    Google Scholar 

  67. Smart O.S., Breed J., Smitgh G.R. and Samsom M.S.P. (1997). A novel method for structure-based prediction of ion channel conductance properties.Biophys. J.72, 1109–1126

    Article  ADS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratory of Membrane Biophysics, Department of Biophysics and Radiobiology, Federal University of Pernambuco, Recife, PE, 50670-901, Brazil

    O. V. Krasilnikov

  2. Laboratory of Molecular Physiology, Institute of Physiology and Biophysics, Tashkent, 700095, Uzbekistan

    O. V. Krasilnikov

Authors
  1. O. V. Krasilnikov
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Biotechnology Division, National Institute of Standards and Technology, Gaithersburg, USA

    John J. Kasianowicz

  2. Department of Biophysics, Pécs University Medical School, Pécs, Hungary

    Miklós S. Z. Kellermayer

  3. Biophysics Laboratory, Department of Chemistry and Biochemistry, University of California, Santa Cruz, USA

    David W. Deamer

Rights and permissions

Reprints and permissions

Copyright information

© 2002 Springer Science+Business Media Dordrecht

About this paper

Cite this paper

Krasilnikov, O.V. (2002). Sizing Channels with Neutral Polymers. In: Kasianowicz, J.J., Kellermayer, M.S.Z., Deamer, D.W. (eds) Structure and Dynamics of Confined Polymers. NATO Science Series, vol 87. Springer, Dordrecht. https://doi.org/10.1007/978-94-010-0401-5_6

Download citation

  • DOI: https://doi.org/10.1007/978-94-010-0401-5_6

  • Publisher Name: Springer, Dordrecht

  • Print ISBN: 978-1-4020-0698-2

  • Online ISBN: 978-94-010-0401-5

  • eBook Packages: Springer Book Archive

Publish with us

Policies and ethics

Search

Navigation