Advertisement

Journal of Biomolecular NMR

, Volume 8, Issue 1, pp 1–14 | Cite as

Macromolecular structural elucidation with solid-state NMR-derived orientational constraints

  • R. R. Ketchem
  • K. -C. Lee
  • S. Huo
  • T. A. Cross
Research Paper

Summary

The complete structure determination of a polypeptide in a lipid bilayer environment is demonstrated built solely upon orientational constraints derived from solid-state NMR observations. Such constraints are obtained from isotopically labeled samples uniformly aligned with respect to the B0 field. Each observation constrains the molecular frame with respect to B0 and the bilayer normal, which are arranged to be parallel. These constraints are not only very precise (a few tenths of a degree), but also very accurate. This is clearly demonstrated as the back bone structure is assembled sequentially and the i to i+6 hydrogen bonds in this structure of the gramicidin channel are shown on average to be within 0.5 Å of ideal geometry. Similarly, the side chains are assembled independently and in a radial direction from the backbone. The lack of considerable atomic overlap between side chains also demonstrates the accuracy of the constraints. Through this complete structure, solid-state NMR is demonstrated as an approach for determining three-dimensional macromolecular structure.

Keywords

Gramicidin A Ion channels 2H NMR Side-chain conformation Membrane protein structure Oriented bilayers 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. BraunW., WiderG., LeeK.H. and WüthrichK. (1983) J. Mol. Biol., 169, 921–948.Google Scholar
  2. BrennemanM.T. and CrossT.A. (1990) J. Chem. Phys., 92, 1483–1494.Google Scholar
  3. Brenneman, M., Quine, J. and Cross, T.A., unpublished results.Google Scholar
  4. CrossT.A. and OpellaS.J. (1983) J. Am. Chem. Soc., 105, 306–308.Google Scholar
  5. CrossT.A., KetchemR.R., HuW., LeeK.-C., LazoN.D. and NorthC.L. (1992) Bull. Magn. Reson., 14, 96–101.Google Scholar
  6. CrossT.A. (1994) Annu. Rep. NMR Spectr., 29, 123–167.Google Scholar
  7. CrossT.A. and OpellaS.J. (1994) Curr. Opin. Struct. Biol., 4, 574–581.Google Scholar
  8. EvansJ.N.S., AppleyardR.J. and ShuttleworthW.A. (1993) J. Am. Chem. Soc., 115, 1588.Google Scholar
  9. FieldsG.B., FieldsC.G., PetefishJ., VanWartH.E. and CrossT.A. (1988) Proc. Natl. Acad. Sci. USA, 85, 1384–1388.Google Scholar
  10. FieldsC.G., FieldsG.B., NobleR.L. and CrossT.A. (1989) Int. J. Pept. Protein Res., 33, 298–303.Google Scholar
  11. GriffithsJ.M. and GriffinR.G. (1993) Anal. Chim. Acta, 283, 1081–1101.Google Scholar
  12. GullionT. and SchaeferJ. (1989) J. Magn. Reson., 81, 196–200.Google Scholar
  13. HarbisonP.K., JelinskiL.W., StarkR.E., TorchiaD.A., HerzfeldJ. and GriffinR.G. (1984) J. Magn. Reson., 60, 79–82.Google Scholar
  14. HartzellC.J., WhitfieldM., OasT.G. and DrobnyG.P. (1987) J. Am. Chem. Soc., 109, 5966–5969.Google Scholar
  15. HendersonR., BaldwinJ.M., CeskaT.A., ZemlinF., BeckmannE. and DowningK.H. (1990) J. Mol. Biol., 213, 899–929.Google Scholar
  16. HuW., LeeK.C. and CrossT.A. (1993) Biochemistry, 32, 7035–7047.Google Scholar
  17. HuW. and CrossT.A. (1995) Biochemistry, 34, 14147–14155.PubMedGoogle Scholar
  18. HuW., LazoN.D. and CrossT.A. (1995) Biochemistry, 34, 14138–14146.Google Scholar
  19. JapB.K., WalianP.J. and GehringK. (1991) Nature, 350, 167–170.Google Scholar
  20. KetchemR.R., HuW. and CrossT.A. (1993) Science, 261, 1457–1460.Google Scholar
  21. Ketchem, R.R. (1995) Structural Determination and Refinement of the Gramicidin A Transmembrane Channel as Studied by Solid-State Nuclear Magnetic Resonance Spectroscopy, Ph.D. Thesis, The Florida State University, Tallahassee, FL.Google Scholar
  22. KetchemR.R., RouxB. and CrossT.A. (1996) In Membrane Structure and Dynamics (Eds, MerzK.M. and RouxB.), Birkhauser, Boston, MA, pp. 299–322.Google Scholar
  23. KuhlbrandtW., WangD. and FujiyoshiY. (1994) Nature, 367, 614–621.Google Scholar
  24. Lazo, N.D., Hu, W. and Cross, T.A. (1992) J. Chem. Soc. Ser. Chem. Commun., 1529–1531.Google Scholar
  25. LazoN.D., HuW., LeeK.C. and CrossT.A. (1993) Biochem. Biophys. Res. Commun., 197, 904–909.Google Scholar
  26. LazoN.D., HuW. and CrossT.A. (1995) J. Magn. Reson., B107, 43–50.Google Scholar
  27. LeeK.C., HuW. and CrossT.A. (1993) Biophys. J., 65, 1162–1167.Google Scholar
  28. LeeK.C., HuoS. and CrossT.A. (1995) Biochemistry, 34, 857–867.Google Scholar
  29. MaiW., HuW., WangC. and CrossT.A. (1993) Protein Sci., 2, 532–542.Google Scholar
  30. McDonnellP.A., ShonK., KimY. and OpellaS.J. (1993) J. Mol. Biol., 233, 447–463.Google Scholar
  31. MichelH. and DeisenhoferJ. (1990) In Current Topics in Membranes and Transport, Vol. 36, Academic Press, New York, NY, pp. 53–69.Google Scholar
  32. MorelliM.A.C., PastoreA. and MottaA. (1992) J. Biomol. NMR, 2, 335.Google Scholar
  33. NorthC.L. and CrossT.A. (1993) J. Magn. Reson., B101, 35–43.Google Scholar
  34. NorthC.L. and CrossT.A. (1995) Biochemistry, 34, 5883–5895.Google Scholar
  35. RoseM.E. (1957) Elementary Theory of Angular Momentum, Wiley, New York, NY.Google Scholar
  36. SchifferM., ChangC.-H. and StevensF.J. (1992) Protein Eng., 5, 213–214.Google Scholar
  37. ShonK., KimY., ColnagoL.A. and OpellaS.J. (1991) Science, 252, 1303–1305.Google Scholar
  38. SmithR. and CornellB.A. (1986) Biophys. J., 49, 117–118.Google Scholar
  39. StarkR.E., JelinskiL.W., RubenD.J., TorchiaD.A. and GriffinR.G. (1983) J. Magn. Reson., 55, 266–273.Google Scholar
  40. TengQ., NicholsonL.K. and CrossT.A. (1991) J. Mol. Biol., 218, 607–619.Google Scholar
  41. TengQ., IqbalM. and CrossT.A. (1992) J. Am. Chem. Soc., 114, 5312–5321.Google Scholar
  42. ValentineK.G., RockwellA.L., GieraschL.M. and OpellaS.J. (1987) J. Magn. Reson., 73, 519–523.Google Scholar
  43. WangD.N. and KuhlbrandtW. (1991) J. Mol. Biol., 217, 691–699.Google Scholar
  44. WeissM.S., AbeleU., WeckesserJ., WelteW., SchiltzE. and SchulzG.E. (1991) Science, 254, 1627–1630.Google Scholar

Copyright information

© ESCOM Science Publishers B.V 1996

Authors and Affiliations

  • R. R. Ketchem
    • 1
    • 2
    • 3
  • K. -C. Lee
    • 1
    • 2
    • 3
  • S. Huo
    • 1
    • 2
    • 3
  • T. A. Cross
    • 1
    • 2
    • 3
  1. 1.Center for Interdisciplinary Magnetic Resonance at the National High Magnetic Field LaboratoryFlorida State UniversityTallahasseeUSA
  2. 2.Institute of Molecular BiophysicsFlorida State UniversityTallahasseeUSA
  3. 3.Department of ChemistryFlorida State UniversityTallahasseeUSA

Personalised recommendations