Skip to main content
SpringerLink
Log in

Quenched Hydrogen Exchange NMR of Amyloid Fibrils

  • Protocol
Protein Amyloid Aggregation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1345))

Abstract

Amyloid fibrils are associated with a number of human diseases. These aggregatively misfolded intermolecular β-sheet assemblies constitute some of the most challenging targets in structural biology because to their complexity, size, and insolubility. Here, protocols and controls are described for experiments designed to study hydrogen-bonding in amyloid fibrils indirectly, by transferring information about amide proton occupancy in the fibrils to the dimethyl sulfoxide-denatured state. Since the denatured state is amenable to solution NMR spectroscopy, the method can provide residue-level-resolution data on hydrogen exchange for the monomers that make up the fibrils.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
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

Abbreviations

DCA:

Dichloroacetic acid

DCA-d2:

Deuterated analog of dichloroacetic acid: Cl2CDCO2D

DCl:

Deuterium chloride (deuterated analog of HCl)

DMSO:

Dimethyl sulfoxide

DMSO-d6:

Deuterated analog of dimethyl sulfoxide (CD3)2SO

D2O:

Deuterium oxide (heavy water)

HSQC:

Heteronuclear single-quantum coherence

IAPP:

Islet amyloid polypeptide

NMR:

Nuclear magnetic resonance

NOESY:

Nuclear Overhauser enhancement spectroscopy

qHX:

Quenched hydrogen exchange

SDS:

Sodium dodecyl sulfate

ThT:

Thioflavin T

TOCSY:

Total correlation spectroscopy

References

  1. Bai Y, Milne JS, Mayne L, Englander SW (1994) Protein stability parameters measured by hydrogen exchange. Proteins 20(1):4–14

    Article  CAS  PubMed  Google Scholar 

  2. Dempsey CE (2001) Hydrogen exchange in peptides and proteins using NMR spectroscopy. Prog Nucl Magn Reson Spectrosc 39:135–170

    Article  CAS  Google Scholar 

  3. Englander SW, Mayne L, Bai Y, Sosnick TR (1997) Hydrogen exchange: the modern legacy of Linderstrom-Lang. Protein Sci 6(5):1101–1109

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  4. Bai Y, Feng H, Zhou Z (2007) Population and structure determination of hidden folding intermediates by native-state hydrogen exchange-directed protein engineering and nuclear magnetic resonance. Methods Mol Biol 350:69–81

    CAS  PubMed  Google Scholar 

  5. Paterson Y, Englander SW, Roder H (1990) An antibody binding site on cytochrome c defined by hydrogen exchange and two-dimensional NMR. Science 249(4970):755–759

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  6. Baldwin RL (2008) The search for folding intermediates and the mechanism of protein folding. Annu Rev Biophys 37:1–21

    Article  CAS  PubMed  Google Scholar 

  7. Roder H, Wuthrich K (1986) Protein folding kinetics by combined use of rapid mixing techniques and NMR observation of individual amide protons. Proteins 1(1):34–42

    Article  CAS  PubMed  Google Scholar 

  8. Alexandrescu AT, Evans PA, Pitkeathly M, Baum J, Dobson CM (1993) Structure and dynamics of the acid-denatured molten globule state of alpha-lactalbumin: a two-dimensional NMR study. Biochemistry 32(7):1707–1718

    Article  CAS  PubMed  Google Scholar 

  9. Alexandrescu AT (2001) An NMR-based quenched hydrogen exchange investigation of model amyloid fibrils formed by cold shock protein A. Pac Symp Biocomput 2001:67–78

    Google Scholar 

  10. Alexandrescu AT, Jahnke W, Wiltscheck R, Blommers MJ (1996) Accretion of structure in staphylococcal nuclease: an 15N NMR relaxation study. J Mol Biol 260(4):570–587

    Article  CAS  PubMed  Google Scholar 

  11. Alexandrescu AT, Abeygunawardana C, Shortle D (1994) Structure and dynamics of a denatured 131-residue fragment of staphylococcal nuclease: a heteronuclear NMR study. Biochemistry 33(5):1063–1072

    Article  CAS  PubMed  Google Scholar 

  12. Santos NC, Figueira-Coelho J, Martins-Silva J, Saldanha C (2003) Multidisciplinary utilization of dimethyl sulfoxide: pharmacological, cellular, and molecular aspects. Biochem Pharmacol 65(7):1035–1041

    Article  CAS  PubMed  Google Scholar 

  13. Voets IK, Cruz WA, Moitzi C, Lindner P, Areas EP, Schurtenberger P (2010) DMSO-induced denaturation of hen egg white lysozyme. J Phys Chem B 114(36):11875–11883

    Article  CAS  PubMed  Google Scholar 

  14. Masters CL, Selkoe DJ (2012) Biochemistry of amyloid beta-protein and amyloid deposits in Alzheimer disease. Cold Spring Harb Perspect Med 2(6):a006262

    Article  PubMed Central  PubMed  Google Scholar 

  15. Zhang YZ, Paterson Y, Roder H (1995) Rapid amide proton exchange rates in peptides and proteins measured by solvent quenching and two-dimensional NMR. Protein Sci 4(4):804–814

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  16. Chandak MS, Nakamura T, Makabe K, Takenaka T, Mukaiyama A, Chaudhuri TK, Kato K, Kuwajima K (2013) The H/D-exchange kinetics of the Escherichia coli co-chaperonin GroES studied by 2D NMR and DMSO-quenched exchange methods. J Mol Biol 425(14):2541–2560

    Article  CAS  PubMed  Google Scholar 

  17. Czerski L, Vinogradova O, Sanders CR (2000) NMR-Based amide hydrogen-deuterium exchange measurements for complex membrane proteins: development and critical evaluation. J Magn Reson 142(1):111–119

    Article  CAS  PubMed  Google Scholar 

  18. Knowles TP, Vendruscolo M, Dobson CM (2014) The amyloid state and its association with protein misfolding diseases. Nat Rev Mol Cell Biol 15(6):384–396

    Article  CAS  PubMed  Google Scholar 

  19. Hoshino M, Katou H, Hagihara Y, Hasegawa K, Naiki H, Goto Y (2002) Mapping the core of the beta(2)-microglobulin amyloid fibril by H/D exchange. Nat Struct Biol 9(5):332–336

    Article  CAS  PubMed  Google Scholar 

  20. Damo SM, Phillips AH, Young AL, Li S, Woods VL Jr, Wemmer DE (2010) Probing the conformation of a prion protein fibril with hydrogen exchange. J Biol Chem 285(42):32303–32311

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. Morgan GJ, Giannini S, Hounslow AM, Craven CJ, Zerovnik E, Turk V, Waltho JP, Staniforth RA (2008) Exclusion of the native alpha-helix from the amyloid fibrils of a mixed alpha/beta protein. J Mol Biol 375(2):487–498

    Article  CAS  PubMed  Google Scholar 

  22. Vilar M, Chou HT, Luhrs T, Maji SK, Riek-Loher D, Verel R, Manning G, Stahlberg H, Riek R (2008) The fold of alpha-synuclein fibrils. Proc Natl Acad Sci U S A 105(25):8637–8642

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  23. Luhrs T, Ritter C, Adrian M, Riek-Loher D, Bohrmann B, Dobeli H, Schubert D, Riek R (2005) 3D structure of Alzheimer’s amyloid-beta(1-42) fibrils. Proc Natl Acad Sci U S A 102(48):17342–17347

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  24. Alexandrescu AT (2013) Amide proton solvent protection in amylin fibrils probed by quenched hydrogen exchange NMR. PLoS One 8(2):e56467

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  25. Hoshino M, Katou H, Yamaguchi K, Goto Y (2007) Dimethylsulfoxide-quenched hydrogen/deuterium exchange method to study amyloid fibril structure. Biochim Biophys Acta 1768(8):1886–1899

    Article  CAS  PubMed  Google Scholar 

  26. Maji SK, Wang L, Greenwald J, Riek R (2009) Structure-activity relationship of amyloid fibrils. FEBS Lett 583(16):2610–2617

    Article  CAS  PubMed  Google Scholar 

  27. Kheterpal I, Wetzel R (2006) Hydrogen/deuterium exchange mass spectrometry – a window into amyloid structure. Acc Chem Res 39(9):584–593

    Article  CAS  PubMed  Google Scholar 

  28. Lee YH, Goto Y (2012) Kinetic intermediates of amyloid fibrillation studied by hydrogen exchange methods with nuclear magnetic resonance. Biochim Biophys Acta 1824(12):1307–1323

    Article  CAS  PubMed  Google Scholar 

  29. Hou L, Shao H, Zhang Y, Li H, Menon NK, Neuhaus EB, Brewer JM, Byeon IJ, Ray DG, Vitek MP, Iwashita T, Makula RA, Przybyla AB, Zagorski MG (2004) Solution NMR studies of the A beta(1-40) and A beta(1-42) peptides establish that the Met35 oxidation state affects the mechanism of amyloid formation. J Am Chem Soc 126(7):1992–2005

    Article  CAS  PubMed  Google Scholar 

  30. Whittemore NA, Mishra R, Kheterpal I, Williams AD, Wetzel R, Serpersu EH (2005) Hydrogen-deuterium (H/D) exchange mapping of Abeta 1-40 amyloid fibril secondary structure using nuclear magnetic resonance spectroscopy. Biochemistry 44(11):4434–4441

    Article  CAS  PubMed  Google Scholar 

  31. Cavanagh J, Fairbrother WJ, Palmer AG III, Rance M, Skelton NJ (2006) Protein NMR spectroscopy principles and practice, 2nd edn. Elsevier Inc., Amsterdam

    Google Scholar 

  32. Olofsson A, Lindhagen-Persson M, Sauer-Eriksson AE, Ohman A (2007) Amide solvent protection analysis demonstrates that amyloid-beta(1-40) and amyloid-beta(1-42) form different fibrillar structures under identical conditions. Biochem J 404(1):63–70

    Article  PubMed Central  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by Basic Research Award 1-10-BS-04 to A.T.A. from the American Diabetes Association. I thank Rebecca L. Newcomer and Anne R. Kaplan for useful comments on the manuscript.

Author information

Authors and Affiliations

  1. Department of Molecular and Cell Biology, University of Connecticut, Unit 3125, BSP 209, 91 N. Eagleville Rd., Storrs, CT, 06269-3125, USA

    Andrei T. Alexandrescu

Authors
  1. Andrei T. Alexandrescu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Andrei T. Alexandrescu .

Editor information

Editors and Affiliations

  1. Weill Cornell Medical College, New York, New York, USA

    David Eliezer

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Alexandrescu, A.T. (2016). Quenched Hydrogen Exchange NMR of Amyloid Fibrils. In: Eliezer, D. (eds) Protein Amyloid Aggregation. Methods in Molecular Biology, vol 1345. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2978-8_14

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-2978-8_14

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2977-1

  • Online ISBN: 978-1-4939-2978-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation