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Journal of Biomolecular NMR

, Volume 44, Issue 4, pp 225–233 | Cite as

Histidine side-chain dynamics and protonation monitored by 13C CPMG NMR relaxation dispersion

  • Mathias A. S. Hass
  • Ali Yilmaz
  • Hans E. M. Christensen
  • Jens J. Led
Article

Abstract

The use of 13C NMR relaxation dispersion experiments to monitor micro-millisecond fluctuations in the protonation states of histidine residues in proteins is investigated. To illustrate the approach, measurements on three specifically 13C labeled histidine residues in plastocyanin (PCu) from Anabaena variabilis (A.v.) are presented. Significant Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion is observed for 13Cε1 nuclei in the histidine imidazole rings of A.v. PCu. The chemical shift changes obtained from the CPMG dispersion data are in good agreement with those obtained from the chemical shift titration experiments, and the CPMG derived exchange rates agree with those obtained previously from 15N backbone relaxation measurements. Compared to measurements of backbone nuclei, 13Cε1 dispersion provides a more direct method to monitor interchanging protonation states or other kinds of conformational changes of histidine side chains or their environment. Advantages and shortcomings of using the 13Cε1 dispersion experiments in combination with chemical shift titration experiments to obtain information on exchange dynamics of the histidine side chains are discussed.

Keywords

Plastocyanin Histidine side-chain dynamics 13C CPMG NMR relaxation dispersion 

Abbreviations

CPMG

Carr-Purcell-Meiboom-Gill

PCu

Plastocyanin

Notes

Acknowledgments

We thank Lise-Lotte Jespersen for technical assistance. This study was supported by the Danish Agency for Science, Technology and Innovation, grants, 21-04-0519 and 272-07-0466, Carlsbergfondet grant 1624/40, Novo Nordisk Fonden grant 2003-11-28, and Villum Kann Rasmussen Fonden grant 8.12.2003.

References

  1. Badsberg U, Jørgensen AMM, Gesmar H, Led JJ, Hammerstad JM, Jespersen LL, Ulstrup J (1996) Solution structure of reduced plastocyanin from the blue-green alga Anabaena variabilis. Biochemistry 35:7021–7031CrossRefGoogle Scholar
  2. Day RM, Thalhauser CJ, Sudmeier JL, Vincent MP, Torchilin EV, Sanford DG, Bachovchin CW, Bachovchin WW (2003) Tautomerism, acid-base equilibria, and H-bonding of the six histidines in subtilisin BPN by NMR. Protein Sci 12:794–810CrossRefGoogle Scholar
  3. Eigen M (1963) Protonenübertragung, Saure-Base-Katalyse und enzymatische hydrolyse. 1. Elementarvorgange. Angew Chem 75:489–508CrossRefGoogle Scholar
  4. Fersht A (1998) Structure and mechanism in protein science: a guide to enzyme catalysis and protein folding. W. H. Freeman and Company, New YorkGoogle Scholar
  5. Grey MJ, Tang YF, Alexov E, McKnight CJ, Raleigh DP, Palmer AG (2006) Characterizing a partially folded intermediate of the villin headpiece domain under non-denaruring conditions: contribution of His41 to the pH-dependent stability of the N-terminal subdomain. J Mol Biol 355:1078–1094CrossRefGoogle Scholar
  6. Guss JM, Harrowell PR, Murata M, Norris VA, Freeman HC (1986) Crystal-structure analyses of reduced (CuI) poplar plastocyanin at 6 pH values. J Mol Biol 192:361–387CrossRefGoogle Scholar
  7. Hass MAS, Thuesen MH, Christensen HEM, Led JJ (2004) Characterization of μs-ms dynamics of proteins using a combined analysis of N-15 NMR relaxation and chemical shift: conformational exchange in plastocyanin induced by histidine protonations. J Am Chem Soc 126:753–765CrossRefGoogle Scholar
  8. Hass MAS, Christensen HEM, Zhang JD, Led JJ (2007) Kinetics and mechanism of the acid transition of the active site in plastocyanin. Biochemistry 46:14619–14628CrossRefGoogle Scholar
  9. Hass MAS, Hansen DF, Christensen HEM, Led JJ, Kay LE (2008a) Characterization of conformational exchange of a histidine side chain: protonation, rotamerization, and tautomerization of his61 in plastocyanin from Anabaena Variabilis. J Am Chem Soc 130:8460–8470CrossRefGoogle Scholar
  10. Hass MAS, Jensen MR, Led JJ (2008b) Probing electric fields in proteins in solution by NMR spectroscopy. Proteins 72:333–343CrossRefGoogle Scholar
  11. Ishima R, Torchia DA (2003) Extending the range of amide proton relaxation dispersion experiments in proteins using a constant-time relaxation-compensated CPMG approach. J Biomol NMR 25:243–248CrossRefGoogle Scholar
  12. Kovrigin EL, Loria JP (2006) Enzyme dynamics along the reaction coordinate: critical role of a conserved residue. Biochemistry 45:2636–2647CrossRefGoogle Scholar
  13. Loria JP, Rance M, Palmer AG (1999a) A relaxation-compensated Carr-Purcell-Meiboom-Gill sequence for characterizing chemical exchange by NMR spectroscopy. J Am Chem Soc 121:2331–2332CrossRefGoogle Scholar
  14. Loria JP, Rance M, Palmer AG (1999b) A TROSY CPMG sequence for characterizing chemical exchange in large proteins. J Biomol NMR 15:151–155CrossRefGoogle Scholar
  15. Luz Z, Meiboom S (1963) Nuclear magnetic resonance study of protolysis of trimethylammonium ion in aqueous solution - order of reaction with respect to solvent. J Chem Phys 39:366–370CrossRefADSGoogle Scholar
  16. Mandel M (1965) Proton magnetic resonance spectra of some proteins. I. Ribonuclease oxidized ribonuclease lysozyme and cytochrome c. J Biol Chem 240:1586–1592Google Scholar
  17. Markley JL (1975) Observation of histidine residues in proteins by means of nuclear magnetic-resonance spectroscopy. Accounts Chem Res 8:70–80CrossRefGoogle Scholar
  18. Palmer AG, Kroenke CD, Loria JP (2001) Nuclear magnetic resonance methods for quantifying microsecond-to-millisecond motions in biological macromolecules. Method Enzymol 339:204–238CrossRefGoogle Scholar
  19. Pelton JG, Torchia DA, Meadow ND, Roseman S (1993) Tautomeric states of the active-site histidines of phosphorylated and unphosphorylated III(Glc), a signal-transducing protein from Escherichia coli, using two-dimensional heteronuclear NMR techniques. Protein Sci 2:543–558CrossRefGoogle Scholar
  20. Perez-Canadilllas JM, Garcia-Mayoral MF, Laurents DV, del Pozo AM, Gavilanes JG, Rico M, Bruix M (2003) Tautomeric state of a-sarcin histidines. Nδ tautomers are a common feature in the active site of extracellular microbial ribonucleases. FEBS Lett 534:197–201CrossRefGoogle Scholar
  21. Reynolds WF, Peat IR, Freedman MH, Lyerla JR (1973) Determination of tautomeric form of imidazole ring of l-histidine in basic solution by C-13 magnetic-resonance spectroscopy. J Am Chem Soc 95:328–331CrossRefGoogle Scholar
  22. Schmidt L, Christensen HEM, Harris P (2006) Structure of plastocyanin from the cyanobacterium Anabaena variabilis. Acta Crystallogr D 62:1022–1029Google Scholar
  23. Shimahara H, Yoshida T, Shibata Y, Shimizu M, Kyogoku Y, Sakiyama F, Nakazawa T, Tate S, Ohki S, Kato T, Moriyama H, Kishida K, Tano Y, Ohkubo T, Kobayashi Y, Ha (2007) Tautomerism of histidine 64 associated with proton transfer in catalysis of carbonic anhydrase. J Biol Chem 282:9646–9656CrossRefGoogle Scholar
  24. Sudmeier JL, Evelhoch JL, Jonsson NBH (1980) Dependence of NMR lineshape analysis upon chemical rates and mechanisms - implications for enzyme histidine titrations. J Magn Reson 40:377–390Google Scholar
  25. Sudmeier JL, Bradshaw EM, Haddad KEC, Day RM, Thalhauser CJ, Bullock PA, Bachovchin WW (2003) Identification of histidine tautomers in proteins by 2D H-1/C-13(δ2) one-bond correlated NMR. J Am Chem Soc 125:8430–8431CrossRefGoogle Scholar
  26. Teilum K, Brath U, Lundstrom P, Akke M (2006) Biosynthetic C-13 labeling of aromatic side chains in proteins for NMR relaxation measurements. J Am Chem Soc 128:2506–2507CrossRefGoogle Scholar
  27. Tollinger M, Skrynnikov NR, Mulder FAA, Forman-Kay JD, Kay LE (2001) Slow dynamics in folded and unfolded states of an SH3 domain. J Am Chem Soc 123:11341–11352CrossRefGoogle Scholar
  28. Vuister GW, Bax A (1992) Resolution enhancement and spectral editing of uniformly C-13-enriched proteins by homonuclear broad-band C-13 decoupling. J Magn Reson 98:428–435Google Scholar

Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  • Mathias A. S. Hass
    • 1
    • 3
  • Ali Yilmaz
    • 1
    • 4
  • Hans E. M. Christensen
    • 2
  • Jens J. Led
    • 1
  1. 1.Department of ChemistryUniversity of CopenhagenCopenhagen ØDenmark
  2. 2.Department of ChemistryThe Technical University of DenmarkLyngbyDenmark
  3. 3.Institute of ChemistryLeiden UniversityLeidenThe Netherlands
  4. 4.Department of Medicinal Chemistry, Faculty of Pharmaceutical SciencesUniversity of CopenhagenCopenhagen ØDenmark

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