Skip to main content
SpringerLink
Log in

Insoluble Protein Assemblies Characterized by Fourier Transform Infrared Spectroscopy

  • Protocol
  • First Online:
Insoluble Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1258))

Abstract

Fourier transform infrared (FTIR) spectroscopy is a useful tool for the structural characterization of insoluble protein assemblies, as it allows to obtain information on the protein secondary structures and on their intermolecular interactions. The protocols for FTIR spectroscopy and microspectroscopy measurements in transmission and attenuated total reflection modes will be presented and illustrated in the following examples: bacterial inclusion bodies, self-assembling peptides, thermal aggregates, and amyloid 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 89.00
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 169.99
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

References

  1. Fink AL (1998) Protein aggregation: folding aggregates, inclusion bodies and amyloid. Fold Des 3:R9–R23

    Article  CAS  PubMed  Google Scholar 

  2. Chiti F, Dobson CM (2006) Protein misfolding, functional amyloid, and human disease. Annu Rev Biochem 75:333–366

    Article  CAS  PubMed  Google Scholar 

  3. Adamcik J, Mezzenga R (2012) Proteins fibrils from a polymer physics perspective. Macromolecules 45:1137–1150

    Article  CAS  Google Scholar 

  4. Cinar G, Ceylan H, Urel M et al (2012) Amyloid inspired self-assembled peptide nanofibers. Biomacromolecules 13:3377–3387

    Article  CAS  PubMed  Google Scholar 

  5. Jonker AM, Lowik DWPM, van Hest JCM (2012) Peptide- and protein-based hydrogels. Chem Mater 24:759–773

    Article  CAS  Google Scholar 

  6. Mezzenga R, Schurtenberger P, Burbidge A et al (2005) Understanding foods as soft materials. Nat Mater 4:729–740

    Article  CAS  PubMed  Google Scholar 

  7. Garcia-Fruitos E, Vazquez E, Diez-Gil C et al (2012) Bacterial inclusion bodies: making gold from waste. Trends Biotechnol 30:65–70

    Article  CAS  PubMed  Google Scholar 

  8. Gatti-Lafranconi P, Natalello A, Ami D et al (2011) Concepts and tools to exploit the potential of bacterial inclusion bodies in protein science and biotechnology. FEBS J 278:2408–2418

    Article  CAS  PubMed  Google Scholar 

  9. Garcia-Fruitos E, Seras-Franzoso J, Vazquez E et al (2010) Tunable geometry of bacterial inclusion bodies as substrate materials for tissue engineering. Nanotechnology 21:205101

    Article  PubMed  Google Scholar 

  10. de Groot NS, Sabate R, Ventura S (2009) Amyloids in bacterial inclusion bodies. Trends Biochem Sci 34:408–416

    Article  PubMed  Google Scholar 

  11. Garcia-Fruitos E, Gonzalez-Montalban N, Morell M et al (2005) Aggregation as bacterial inclusion bodies does not imply inactivation of enzymes and fluorescent proteins. Microb Cell Fact 4:27

    Article  PubMed Central  PubMed  Google Scholar 

  12. Seshadri S, Khurana R, Fink AL (1999) Fourier transform infrared spectroscopy in analysis of protein deposits. Methods Enzymol 309:559–576

    Article  CAS  PubMed  Google Scholar 

  13. Ami D, Natalello A, Doglia SM (2012) Fourier transform infrared microspectroscopy of complex biological systems: from intact cells to whole organisms. In: Uversky VN, Dunker AK (eds) Intrinsically disordered protein analysis: volume 1, vol 895, Methods and experimental tools. Methods in molecular biology. Humana, New York, pp 85–100. doi:10.1007/978-1-61779-927-3_7

    Chapter  Google Scholar 

  14. Natalello A, Ami D, Doglia S (2012) Fourier transform infrared spectroscopy of intrinsically disordered proteins: measurement procedures and data analyses. In: Uversky VN, Dunker AK (eds) Intrinsically disordered protein analysis: volume 1, methods and experimental tools. Methods in molecular biology, vol 895. Humana, New York, pp 229–244. doi:10.1007/978-1-61779-927-3_16

  15. Barth A (2007) Infrared spectroscopy of proteins. Biochim Biophys Acta 1767:1073–1101

    Article  CAS  PubMed  Google Scholar 

  16. Zandomeneghi G, Krebs MRH, McCammon MG et al (2004) FTIR reveals structural differences between native beta-sheet proteins and amyloid fibrils. Protein Sci 13:3314–3321

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  17. Natalello A, Ami D, Brocca S et al (2005) Secondary structure, conformational stability and glycosylation of a recombinant Candida rugosa lipase studied by Fourier-transform infrared spectroscopy. Biochem J 385:511–517

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  18. Natalello A, Doglia SM, Carey J et al (2007) Role of flavin mononucleotide in the thermostability and oligomerization of Escherichia coli stress-defense protein WrbA. Biochemistry 46:543–553

    Article  CAS  PubMed  Google Scholar 

  19. Natalello A, Frana AM, Relini A et al (2011) A major role for side-chain polyglutamine hydrogen bonding in irreversible ataxin-3 aggregation. PLoS One 6:e18789

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  20. Goormaghtigh E, Raussens V, Ruysschaert JM (1999) Attenuated total reflection infrared spectroscopy of proteins and lipids in biological membranes. Biochim Biophys Acta 1422:105–185

    Article  CAS  PubMed  Google Scholar 

  21. Doglia SM, Ami D, Natalello A et al (2008) Fourier transform infrared spectroscopy analysis of the conformational quality of recombinant proteins within inclusion bodies. Biotechnol J 3:193–201

    Article  CAS  PubMed  Google Scholar 

  22. Carpenter JF, Prestrelski SJ, Dong A (1998) Application of infrared spectroscopy to development of stable lyophilized protein formulations. Eur J Pharm Biopharm 45:231–238

    Article  CAS  PubMed  Google Scholar 

  23. Ami D, Ricagno S, Bolognesi M et al (2012) Structure, stability, and aggregation of β-2 microglobulin mutants: insights from a Fourier transform infrared study in solution and in the crystalline state. Biophys J 102:1676–1684

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  24. Barth A, Zscherp C (2002) What vibrations tell us about proteins. Q Rev Biophys 35:369–430

    Article  CAS  PubMed  Google Scholar 

  25. Arrondo JLR, Goni FM (1999) Structure and dynamics of membrane proteins as studied by infrared spectroscopy. Prog Biophys Mol Biol 72:367–405

    Article  CAS  PubMed  Google Scholar 

  26. Tamm LK, Tatulian SA (1997) Infrared spectroscopy of proteins and peptides in lipid bilayers. Q Rev Biophys 30:365–429

    Article  CAS  PubMed  Google Scholar 

  27. Gelain F, Silva D, Caprini A et al (2011) BMHP1-derived self-assembling peptides: hierarchically assembled structures with self-healing propensity and potential for tissue engineering applications. ACS Nano 5:1845–1859

    Article  CAS  PubMed  Google Scholar 

  28. Silva D, Natalello A, Sanii B et al (2013) Synthesis and characterization of designed BMHP1-derived self-assembling peptides for tissue engineering applications. Nanoscale 5:704–718

    Article  CAS  PubMed  Google Scholar 

  29. Invernizzi G, Aprile FA, Natalello A et al (2012) The relationship between aggregation and toxicity of polyglutamine-containing ataxin-3 in the intracellular environment of Escherichia coli. PLoS One 7:e51890

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  30. Costa MC, Paulson HL (2012) Toward understanding Machado–Joseph disease. Prog Neurobiol 97:239–257

    Article  CAS  PubMed Central  Google Scholar 

  31. Rahmelow K, Hubner W (1997) Infrared spectroscopy in aqueous solution: difficulties and accuracy of water subtraction. Appl Spectrosc 51:160–170

    Article  CAS  Google Scholar 

  32. Venyaminov SY, Prendergast FG (1997) Water (H2O and D2O) molar absorptivity in the 1000–4000 cm−1 range and quantitative infrared spectroscopy of aqueous solutions. Anal Biochem 248:234–245

    Article  CAS  PubMed  Google Scholar 

  33. Dong A, Huang P, Caughey WS (1990) Protein secondary structures in water from 2nd-derivative amide-I infrared-spectra. Biochemistry 29:3303–3308

    Article  CAS  PubMed  Google Scholar 

  34. Natalello A, Prokorov VV, Tagliavini F et al (2008) Conformational plasticity of the Gerstmann-Straussler-Scheinker disease peptide as indicated by its multiple aggregation pathways. J Mol Biol 381:1349–1361

    Article  CAS  PubMed  Google Scholar 

  35. Ami D, Natalello A, Lotti M et al (2013) Why and how protein aggregation has to be studied in vivo. Microb Cell Fact 12:17

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  36. Choo LP, Wetzel DL, Halliday WC et al (1996) In situ characterization of beta-amyloid in Alzheimer’s diseased tissue by synchrotron Fourier transform infrared microspectroscopy. Biophys J 71:1672–1679

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  37. Diomede L, Cassata G, Fiordaliso F et al (2010) Tetracycline and its analogues protect Caenorhabditis elegans from beta amyloid-induced toxicity by targeting oligomers. Neurobiol Dis 40:424–431

    Article  CAS  PubMed  Google Scholar 

  38. Ami D, Bonecchi L, Cali S et al (2003) FT-IR study of heterologous protein expression in recombinant Escherichia coli strains. Biochim Biophys Acta 1624:6–10

    Article  CAS  PubMed  Google Scholar 

  39. Ami D, Natalello A, Gatti-Lafranconi P et al (2005) Kinetics of inclusion body formation studied in intact cells by FT-IR spectroscopy. FEBS Lett 579:3433–3436

    Article  CAS  PubMed  Google Scholar 

  40. Ami D, Natalello A, Taylor G et al (2006) Structural analysis of protein inclusion bodies by Fourier transform infrared microspectroscopy. Biochim Biophys Acta 1764:793–799

    Article  CAS  PubMed  Google Scholar 

  41. Gonzalez-Montalban N, Natalello A, Garcia-Fruitos E et al (2008) In situ protein folding and activation in bacterial inclusion bodies. Biotechnol Bioeng 100:797–802

    Article  CAS  PubMed  Google Scholar 

  42. Susi H, Byler DM (1986) Resolution-enhanced Fourier-transform infrared-spectroscopy of enzymes. Methods Enzymol 130:290–311

    Article  CAS  PubMed  Google Scholar 

  43. Arrondo JLR, Muga A, Castresana J et al (1993) Quantitative studies of the structure of proteins in solution by Fourier-transform infrared-spectroscopy. Prog Biophys Mol Biol 59:23–25

    Article  CAS  PubMed  Google Scholar 

  44. Kauppinen JK, Moffatt DJ, Mantsch HH et al (1981) Fourier self-deconvolution – a method for resolving intrinsically overlapped bands. Appl Spectrosc 35:271–276

    Article  CAS  Google Scholar 

  45. Goormaghtigh E, Cabiaux V, Ruysschaert JM (1994) Determination of soluble and membrane protein structure by Fourier transform infrared spectroscopy. III. Secondary structures. Sub-cell Biochem 23:405–450

    Article  CAS  Google Scholar 

  46. Khurana R, Fink AL (2000) Do parallel beta-helix proteins have a unique Fourier transform infrared spectrum? Biophys J 78:994–1000

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  47. Susi H, Byler DM (1987) Fourier-transform infrared study of proteins with parallel beta-chains. Arch Biochem Biophys 258:465–469

    Article  CAS  PubMed  Google Scholar 

  48. Chitnumsub P, Fiori WR, Lashuel HA et al (1999) The nucleation of monomeric parallel beta-sheet-like structures and their self-assembly in aqueous solution. Bioorg Med Chem 7:39–59

    Article  CAS  PubMed  Google Scholar 

  49. Yamada N, Ariga K, Naito M et al (1998) Regulation of b-sheet structures within amyloid-like b-sheet assemblage from tripeptide derivatives. J Am Chem Soc 120:12192–12199

    Article  CAS  Google Scholar 

  50. Cerf E, Sarroukh R, Tamamizu‑Kato S et al (2009) Antiparallel β-sheet: a signature structure of the oligomeric amyloid β-peptide. Biochem J 421:415–423

    Article  CAS  PubMed  Google Scholar 

  51. Sarroukh R, Goormaghtigh E, Ruysschaert JM et al (2013) ATR-FTIR: a “rejuvenated” tool to investigate amyloid proteins. Biochim Biophys Acta 1828:2328–2338

    Article  CAS  PubMed  Google Scholar 

  52. Torii H, Tatsumi T, Tasumi M (1998) Effects of hydration on the structure, vibrational wavenumbers, vibrational force field and resonance raman intensities of N-methylacetamide. J Raman Spectrosc 29:537–546

    Article  CAS  Google Scholar 

  53. Natalello A, Mattoo RUH, Priya S et al (2013) Biophysical characterization of two different stable misfolded monomeric polypeptides that are chaperone-amenable substrates. J Mol Biol 425:1158–1171

    Article  CAS  PubMed  Google Scholar 

  54. Kauffmann E, Darnton NC, Austin RH et al (2001) Lifetimes of intermediates in the beta-sheet to alpha-helix transition of beta-lactoglobulin by using a diffusional IR mixer. Proc Natl Acad Sci U S A 98:6646–6649

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  55. Tatulian SA, Biltonen RL, Tamm LK (1997) Structural changes in a secretory phospholipase A(2) induced by membrane binding: a clue to interfacial activation? J Mol Biol 268:809–815

    Article  CAS  PubMed  Google Scholar 

  56. Walsh STR, Cheng RP, Wright WW et al (2003) The hydration of amides in helices; a comprehensive picture from molecular dynamics, IR, and NMR. Protein Sci 12:520–531

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  57. Martinez G, Millhauser G (1995) Ftir spectroscopy of alanine-based peptides – assignment of the amide I' modes for random coil and helix. J Struct Biol 114:23–27

    Article  CAS  PubMed  Google Scholar 

  58. Javor S, Natalello A, Doglia SM et al (2008) Alpha-helix stabilization within a peptide dendrimer. J Am Chem Soc 130:17248–17249

    Article  CAS  PubMed  Google Scholar 

  59. Roque A, Iloro I, Ponte I et al (2005) DNA-induced secondary structure of the carboxyl-terminal domain of histone H1. J Biol Chem 280:32141–32147

    Article  CAS  PubMed  Google Scholar 

  60. Barth A (2000) The infrared absorption of amino acid side chains. Prog Biophys Mol Biol 74:141–173

    Article  CAS  PubMed  Google Scholar 

  61. Haris PI, Severcan F (1999) FTIR spectroscopic characterization of protein structure in aqueous and non-aqueous media. J Mol Catal B: Enzym 7:207–221

    Article  CAS  Google Scholar 

  62. Hering JA, Innocent PR, Haris PI (2002) Automatic amide I frequency selection for rapid quantification of protein secondary structure from Fourier transform infrared spectra of proteins. Proteomics 2:839–849

    Article  CAS  PubMed  Google Scholar 

  63. Vila R, Ponte I, Collado M et al (2001) Induction of secondary structure in a COOH-terminal peptide of histone H1 by interaction with the DNA. J Biol Chem 276:30898–30903

    Article  CAS  PubMed  Google Scholar 

  64. Salomaa P, Schaleger LL, Long FA (1964) Solvent deuterium isotope effects on acid–base equilibria. J Am Chem Soc 86:1–7

    Article  CAS  Google Scholar 

  65. Dong AC, Hyslop RM, Pringle DL (1996) Differences in conformational dynamics of ribonucleases A and S as observed by infrared spectroscopy and hydrogen-deuterium exchange. Arch Biochem Biophys 333:275–281

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgment

In this work to illustrate the protocols for FTIR measurements of protein aggregates, we presented examples on proteins studied in collaborations with colleagues that we would like to acknowledge: Prof. Paolo Tortora, Prof. Angelo Vescovi, Dr. Diletta Ami, Dr. Maria E. Regonesi, and Dr. Fabrizio Gelain of the University of Milano-Bicocca, Prof. Martino Bolognesi and Dr. Stefano Ricagno of the University of Milan, and Prof. Vittorio Bellotti of the University of Pavia.

Author information

Authors and Affiliations

  1. Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, Milan, 20126, Italy

    Antonino Natalello & Silvia M. Doglia

  2. Department of Physics, University of Milano-Bicocca, Piazza della Scienza 3, 20126, Milan, Italy

    Antonino Natalello & Silvia M. Doglia

  3. Consorzio Nazionale Interuniversitario per le Scienze Fisiche della Materia (CNISM), UdR Milano-Bicocca, Milan, Italy

    Antonino Natalello & Silvia M. Doglia

Authors
  1. Antonino Natalello
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Silvia M. Doglia
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Antonino Natalello .

Editor information

Editors and Affiliations

  1. Departament de Genètica i de Microbiologia, CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN) and Universitat Autònoma de Barcelona, Bellaterra (Barcelona), Spain

    Elena García-Fruitós

Rights and permissions

Reprints and permissions

Copyright information

© 2015 Springer Science+Business Media New York

About this protocol

Cite this protocol

Natalello, A., Doglia, S.M. (2015). Insoluble Protein Assemblies Characterized by Fourier Transform Infrared Spectroscopy. In: García-Fruitós, E. (eds) Insoluble Proteins. Methods in Molecular Biology, vol 1258. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2205-5_20

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-2205-5_20

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2204-8

  • Online ISBN: 978-1-4939-2205-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation