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Isotope Labeling Methods for Large Systems

  • Patrik LundströmEmail author
  • Alexandra Ahlner
  • Annica Theresia Blissing
Chapter
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 992)

Abstract

A major drawback of nuclear magnetic resonance (NMR) spectroscopy compared to other methods is that the technique has been limited to relatively small molecules. However, in the last two decades the size limit has been pushed upwards considerably and it is now possible to use NMR spectroscopy for structure calculations of proteins of molecular weights approaching 100 kDa and to probe dynamics for supramolecular complexes of molecular weights in excess of 500 kDa. Instrumental for this progress has been development in instrumentation and pulse sequence design but also improved isotopic labeling schemes that lead to increased sensitivity as well as improved spectral resolution and simplification. These are described and discussed in this chapter, focusing on labeling schemes for amide proton and methyl proton detected experiments. We also discuss labeling methods for other potentially useful positions in proteins.

Keywords

Nuclear Magnetic Resonance Nuclear Magnetic Resonance Spectroscopy Amide Proton Nuclear Overhauser Effect Residual Dipolar Coupling 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. 1.
    Purcell EM, Torrey HC, Pound RV (1946) Resonance absorption of nuclear magnetic moments in a solid. Phys Rev 69:37–38CrossRefGoogle Scholar
  2. 2.
    Bloch F, Hansen WW, Packard ME (1946) Nuclear induction. Phys Rev 69:680Google Scholar
  3. 3.
    Proctor WG, Yu FC (1950) The dependence of a nuclear magnetic resonance frequency upon chemical compounds. Phys Rev 77:717CrossRefGoogle Scholar
  4. 4.
    Dickinson WC (1950) Dependence of the 19F nuclear resonance position on chemical compound. Phys Rev 77:736–737CrossRefGoogle Scholar
  5. 5.
    Overhauser AW (1953) Polarization of nuclei in materials. Phys Rev 92:411–415CrossRefGoogle Scholar
  6. 6.
    Carver TR, Slichter CP (1953) Polarization of nuclear spins in metals. Phys Rev 92:212–213CrossRefGoogle Scholar
  7. 7.
    Solomon I (1955) Relaxation processes in a system of two spins. Phys Rev 99:559–566CrossRefGoogle Scholar
  8. 8.
    Wagner G, Wüthrich K (1982) Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra: basic pancreatic trypsin inhibitor. J Mol Biol 155:347–366PubMedCrossRefGoogle Scholar
  9. 9.
    Williamson MP, Havel TF, Wüthrich K (1985) Solution conformation of proteinase inhibitor IIa from bull seminal plasma by 1H nuclear magnetic resonance and distance geometry. J Mol Biol 182:295–315PubMedCrossRefGoogle Scholar
  10. 10.
    Oschkinat H, Griesinger C, Kraulis PJ, Sorensen OW, Ernst RR, Gronenborn AM, Clore GM (1988) 3-dimensional NMR spectroscopy of a protein in solution. Nature 332:374–376PubMedCrossRefGoogle Scholar
  11. 11.
    Fesik SW, Zuiderweg ERP (1988) Heteronuclear 3-dimensional NMR spectroscopy – a strategy for the simplification of homonuclear two-dimensional NMR spectra. J Magn Reson 78:588–593Google Scholar
  12. 12.
    Marion D, Driscoll PC, Kay LE, Wingfield PT, Bax A, Gronenborn AM, Clore GM (1989) Overcoming the overlap problem in the assignment of 1H NMR spectra of larger proteins by use of 3-dimensional heteronuclear 1H-15N Hartmann-Hahn multiple quantum coherence and nuclear Overhauser multiple quantum coherence spectroscopy – application to interleukin-1 beta. Biochemistry 28:6150–6156PubMedCrossRefGoogle Scholar
  13. 13.
    Ikura M, Kay LE, Tschudin R, Bax A (1990) 3-dimensional NOESY-HMQC spectroscopy of a 13C labeled protein. J Magn Reson 86:204–209Google Scholar
  14. 14.
    Kay LE, Ikura M, Tschudin R, Bax A (1990) Three-dimensional triple-resonance NMR spectroscopy of isotopically enriched proteins. J Magn Reson 89:496–514Google Scholar
  15. 15.
    Tjandra N, Bax A (1997) Direct measurement of distances and angles in biomolecules by NMR in a dilute liquid crystalline medium. Science 278:1111–1114PubMedCrossRefGoogle Scholar
  16. 16.
    Pervushin K, Riek R, Wider G, Wüthrich K (1998) Transverse relaxation-optimized spectroscopy (TROSY) for NMR studies of aromatic spin systems in 13C-labeled proteins. J Am Chem Soc 120:6394–6400CrossRefGoogle Scholar
  17. 17.
    Tugarinov V, Muhandiram R, Ayed A, Kay LE (2002) Four-dimensional NMR spectroscopy of a 723-residue protein: chemical shift assignments and secondary structure of malate synthase G. J Am Chem Soc 124:10025–10035PubMedCrossRefGoogle Scholar
  18. 18.
    Goto NK, Gardner KH, Mueller GA, Willis RC, Kay LE (1999) A robust and cost-effective method for the production of Val, Leu, Ile (δ1) methyl-protonated 15N-, 13C-, 2H-labeled proteins. J Biomol NMR 13:369–374PubMedCrossRefGoogle Scholar
  19. 19.
    Tugarinov V, Kay LE (2003) Ile, Leu, and Val methyl assignments of the 723-residue malate synthase G using a new labeling strategy and novel NMR methods. J Am Chem Soc 125:13868–13878PubMedCrossRefGoogle Scholar
  20. 20.
    Pervushin K, Riek R, Wider G, Wüthrich K (1997) Attenuated T2 relaxation by mutual cancellation of dipole-dipole coupling and chemical shift anisotropy indicates an avenue to NMR structures of very large biological macromolecules in solution. Proc Natl Acad Sci USA 94:12366–12371PubMedCrossRefGoogle Scholar
  21. 21.
    Cavanagh J, Fairbrother WJ, Palmer AG 3rd, Rance M, Skelton NJ (2007) Protein NMR spectroscopy: principles and practice. Elsevier Academic Press, BurlingtonGoogle Scholar
  22. 22.
    Lipari G, Szabo A (1982) Model-free approach to the interpretation of nuclear magnetic-resonance relaxation in macromolecules.1. Theory and range of validity. J Am Chem Soc 104:4546–4559CrossRefGoogle Scholar
  23. 23.
    Lipari G, Szabo A (1982) Model-free approach to the interpretation of nuclear magnetic-resonance relaxation in macromolecules. 2. Analysis of experimental results. J Am Chem Soc 104:4559–4570CrossRefGoogle Scholar
  24. 24.
    Clore GM, Szabo A, Bax A, Kay LE, Driscoll PC, Gronenborn AM (1990) Deviations from the simple two-parameter model-free approach to the interpretation of 15N nuclear magnetic relaxation of proteins. J Am Chem Soc 112:4989–4991CrossRefGoogle Scholar
  25. 25.
    Ollerenshaw JE, Tugarinov V, Kay LE (2003) Methyl TROSY: explanation and experimental verification. Magn Reson Chem 41:843–852CrossRefGoogle Scholar
  26. 26.
    Miclet E, Williams DC Jr, Clore GM, Bryce DL, Boisbouvier J, Bax A (2004) Relaxation-optimized NMR spectroscopy of methylene groups in proteins and nucleic acids. J Am Chem Soc 126:10560–10570PubMedCrossRefGoogle Scholar
  27. 27.
    Tugarinov V, Hwang PM, Ollerenshaw JE, Kay LE (2003) Cross-correlated relaxation enhanced H-1-C-13 NMR spectroscopy of methyl groups in very high molecular weight proteins and protein complexes. J Am Chem Soc 125:10420–10428PubMedCrossRefGoogle Scholar
  28. 28.
    Czisch M, Boelens R (1998) Sensitivity enhancement in the TROSY experiment. J Magn Reson 134:158–160PubMedCrossRefGoogle Scholar
  29. 29.
    Weigelt J (1998) Single scan, sensitivity- and gradient-enhanced TROSY for multidimensional NMR experiments. J Am Chem Soc 120:10778–10779CrossRefGoogle Scholar
  30. 30.
    Nietlispach D (2005) Suppression of anti-TROSY lines in a sensitivity enhanced gradient selection TROSY scheme. J Biomol NMR 31:161–166PubMedCrossRefGoogle Scholar
  31. 31.
    Salzmann M, Pervushin K, Wider G, Senn H, Wüthrich K (1998) TROSY in triple-resonance experiments: new perspectives for sequential NMR assignment of large proteins. Proc Natl Acad Sci USA 95:13585–13590PubMedCrossRefGoogle Scholar
  32. 32.
    Yang DW, Kay LE (1999) Improved 1HN-detected triple resonance TROSY-based experiments. J Biomol NMR 13:3–10PubMedCrossRefGoogle Scholar
  33. 33.
    Tugarinov V, Sprangers R, Kay LE (2004) Line narrowing in methyl-TROSY using zero-quantum 1H-13C NMR spectroscopy. J Am Chem Soc 126:4921–4925PubMedCrossRefGoogle Scholar
  34. 34.
    Maniatis T, Sambrook J, Fritsch EF (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, pp 68–69Google Scholar
  35. 35.
    Middelberg APJ (2002) Preparative protein refolding. Trends Biotechnol 20:437–443PubMedCrossRefGoogle Scholar
  36. 36.
    Xu YQ, Zheng Y, Fan JS, Yang DW (2006) A new strategy for structure determination of large proteins in solution without deuteration. Nat Methods 3:931–937PubMedCrossRefGoogle Scholar
  37. 37.
    Bayrhuber M, Riek R (2011) Very simple combination of TROSY, CRINEPT and multiple quantum coherence for signal enhancement in an HN(CO)CA experiment for large proteins. J Magn Reson 209:310–314PubMedCrossRefGoogle Scholar
  38. 38.
    Schanda P, Van Melckebeke H, Brutscher B (2006) Speeding up three-dimensional protein NMR experiments to a few minutes. J Am Chem Soc 128:9042–9043PubMedCrossRefGoogle Scholar
  39. 39.
    Tugarinov V, Choy WY, Orekhov VY, Kay LE (2005) Solution NMR-derived global fold of a monomeric 82-kDa enzyme. Proc Natl Acad Sci USA 102:622–627PubMedCrossRefGoogle Scholar
  40. 40.
    Fiaux J, Bertelsen EB, Horwich AL, Wüthrich K (2002) NMR analysis of a 900 K GroEL GroES complex. Nature 418:207–211PubMedCrossRefGoogle Scholar
  41. 41.
    McIntosh LP, Dahlquist FW (1990) Biosynthetic incorporation of 15N and 13C for assignment and interpretation of nuclear magnetic resonance spectra of proteins. Q Rev Biophys 23:1–38PubMedCrossRefGoogle Scholar
  42. 42.
    Fiaux J, Bertelsen EB, Horwich AL, Wüthrich K (2004) Uniform and residue-specific 15N-labeling of proteins on a highly deuterated background. J Biomol NMR 29:289–297PubMedCrossRefGoogle Scholar
  43. 43.
    Baldwin RL (2002) Making a network of hydrophobic clusters. Science 295:1657–1658PubMedCrossRefGoogle Scholar
  44. 44.
    Bogan AA, Thorn KS (1998) Anatomy of hot spots in protein interfaces. J Mol Biol 280:1–9PubMedCrossRefGoogle Scholar
  45. 45.
    Rodriguez-Mias RA, Pellecchia M (2003) Use of selective Trp side chain labeling to characterize protein-protein and protein-ligand interactions by NMR spectroscopy. J Am Chem Soc 125:2892–2893PubMedCrossRefGoogle Scholar
  46. 46.
    Löhr F, Katsemi V, Betz M, Hartleib J, Rüterjans H (2002) Sequence-specific assignment of histidine and tryptophan ring 1H, 13C and 15N resonances in 13C/15N- and 2H/13C/15N-labelled proteins. J Biomol NMR 22:153–164PubMedCrossRefGoogle Scholar
  47. 47.
    Takahashi H, Nakanishi T, Kami K, Arata Y, Shimada I (2000) A novel NMR method for determining the interfaces of large protein-protein complexes. Nat Struct Biol 7:220–223PubMedCrossRefGoogle Scholar
  48. 48.
    Meissner A, Sorensen OW (1999) Optimization of three-dimensional TROSY-type HCCH NMR correlation of aromatic 1H-13C groups in proteins. J Magn Reson 139:447–450PubMedCrossRefGoogle Scholar
  49. 49.
    Kainosho M, Torizawa T, Iwashita Y, Terauchi T, Ono AM, Guntert P (2006) Optimal isotope labelling for NMR protein structure determinations. Nature 440:52–57PubMedCrossRefGoogle Scholar
  50. 50.
    Ruschak AM, Kay LE (2010) Methyl groups as probes of supra-molecular structure, dynamics and function. J Biomol NMR 46:75–87PubMedCrossRefGoogle Scholar
  51. 51.
    Sprangers R, Kay LE (2007) Quantitative dynamics and binding studies of the 20S proteasome by NMR. Nature 445:618–622PubMedCrossRefGoogle Scholar
  52. 52.
    Gelis I, Bonvin A, Keramisanou D, Koukaki M, Gouridis G, Karamanou S, Economou A, Kalodimos CG (2007) Structural basis for signal-sequence recognition by the translocase motor SecA as determined by NMR. Cell 131:756–769PubMedCrossRefGoogle Scholar
  53. 53.
    Religa TL, Kay LE (2010) Optimal methyl labeling for studies of supra-molecular systems. J Biomol NMR 47:163–169PubMedCrossRefGoogle Scholar
  54. 54.
    Ruschak AM, Velyvis A, Kay LE (2010) A simple strategy for 13C,1H labeling at the Ile-gamma 2 methyl position in highly deuterated proteins. J Biomol NMR 48:129–135PubMedCrossRefGoogle Scholar
  55. 55.
    Religa TL, Sprangers R, Kay LE (2010) Dynamic regulation of archaeal proteasome gate opening as studied by TROSY NMR. Science 328:98–102PubMedCrossRefGoogle Scholar
  56. 56.
    Kigawa T, Muto Y, Yokoyama S (1995) Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for NMR analysis. J Biomol NMR 6:129–134PubMedCrossRefGoogle Scholar
  57. 57.
    Etezady-Esfarjani T, Hiller S, Villalba C, Wüthrich K (2007) Cell-free protein synthesis of perdeuterated proteins for NMR studies. J Biomol NMR 39:229–238PubMedCrossRefGoogle Scholar
  58. 58.
    Goerke AR, Swartz JR (2009) High-level cell-free synthesis yields of proteins containing site-specific non-natural amino acids. Biotechnol Bioeng 102:400–416PubMedCrossRefGoogle Scholar
  59. 59.
    Ruschak AM, Religa TL, Breuer S, Witt S, Kay LE (2010) The proteasome antechamber maintains substrates in an unfolded state. Nature 467:868–871PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2012

Authors and Affiliations

  • Patrik Lundström
    • 1
    Email author
  • Alexandra Ahlner
    • 1
  • Annica Theresia Blissing
    • 1
  1. 1.Division of Molecular Biotechnology, Department of Physics, Chemistry and BiologyLinköping UniversityLinköpingSweden

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