Skip to main content
SpringerLink
Log in

Confocal Fluorescence Detected Linear Dichroism Imaging of Isolated Human Amyloid Fibrils. Role of Supercoiling

  • Original Paper
  • Published:
Journal of Fluorescence Aims and scope Submit manuscript

Abstract

Amyloids are highly organized insoluble protein aggregates that are associated with a large variety of degenerative diseases. In this work, we investigated the anisotropic architecture of isolated human amyloid samples stained with Congo Red. This was performed by fluorescence detected linear dichroism (FDLD) imaging in a laser scanning confocal microscope that was equipped with a differential polarization attachment using high frequency modulation of the polarization state of the laser beam and a demodulation circuit. Two- and three-dimensional FDLD images of amyloids provided information on the orientation of the electric transition dipoles of the intercalated Congo Red molecules with unprecedented precision and spatial resolution. We show that, in accordance with linear dichroism imaging (Jin et al. Proc Natl Acad Sci USA 100:15294, 2003), amyloids exhibit strong anisotropy with preferential orientation of the dye molecules along the fibrils; estimations on the orientation angle, of around 45°, are given using a model calculation which takes into account the helical organization of the filaments and fibrils. Our data also show that FDLD images display large inhomogeneities, high local values with alternating signs and, in some regions, well identifiable µm-sized periodicities. These features of the anisotropic architecture are accounted for by supercoiling of helically organized amyloid fibrils.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  1. Sipe JD, Cohen AS (2000) Review: history of the amyloid fibril. J Struct Biol 130(2–3):88–98

    Article  PubMed  CAS  Google Scholar 

  2. Kelly JW (1998) The alternative conformations of amyloidogenic proteins and their multi-step assembly pathways. Curr Opin Struct Biol 8(1):101–106

    Article  PubMed  CAS  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  4. Dobson CM (2003) Protein folding and misfolding. Nature 426(6968):884–890

    Article  PubMed  CAS  Google Scholar 

  5. Makabe K, Biancalana M, Yan S, Tereshko V, Gawlak G, Miller-Auer H, Meredith SC, Koide S (2008) High-resolution structure of a self-assembly-competent form of a hydrophobic peptide captured in a soluble beta-sheet scaffold. J Mol Biol 378(2):459–467

    Article  PubMed  CAS  Google Scholar 

  6. Karsai A, Martonfalvi Z, Nagy A, Grama L, Penke B, Kellermayer MSZ (2006) Mechanical manipulation of Alzheimer’s amyloid beta 1–42 fibrils. J Struct Biol 155(2):316–326

    Article  PubMed  CAS  Google Scholar 

  7. Giurleo JT, He X, Talaga DS (2008) β-Lactoglobulin assembles into amyloid through sequential aggregated intermediates. J Mol Biol 381(5):1332–1348

    Article  PubMed  CAS  Google Scholar 

  8. Sunde M, Serpell LC, Bartlam M, Fraser PE, Pepys MB, Blake CCF (1997) The common core structure of amyloid fibrils by synchrotron X-ray diffraction. J Mol Biol 273(3):729–739

    Article  PubMed  CAS  Google Scholar 

  9. Blake C, Serpell LC (1996) Synchrotron X-ray studies suggest that the core of the transthyretin amyloid fibril is a continuous beta-sheet helix. Structure 4(8):989–998

    Article  PubMed  CAS  Google Scholar 

  10. Kodali R, Wetzel R (2007) Polymorphism in the intermediates and products of amyloid assembly. Curr Opin Struct Biol 17(1):48–57

    Article  PubMed  CAS  Google Scholar 

  11. Kellermayer MSZ, Karsai A, Benke M, Soos K, Penke B (2008) Stepwise dynamics of epitaxially growing single amyloid fibrils. Proc Natl Acad Sci USA 105(1):141–144

    Article  PubMed  CAS  Google Scholar 

  12. Celej MS, Caarls W, Demchenko AP, Jovin TM (2009) A triple-emission fluorescent probe reveals distinctive amyloid fibrillar polymorphism of wild-type alpha-synuclein and its familial Parkinson’s disease mutants. Biochemistry 11;48(31):7465–7472

    Google Scholar 

  13. Jimenez JL, Guijarro JI, Orlova E, Zurdo J, Dobson CM, Sunde M, Saibil HR (1999) Cryo-electron microscopy structure of an SH3 amyloid fibril and model of the molecular packing. EMBO J 18(4):815–821

    Article  PubMed  CAS  Google Scholar 

  14. Kirschner DA, Abraham C, Selkoe DJ (1986) X-ray diffraction from intraneuronal paired helical filaments and extraneuronal amyloid fibers in Alzheimer disease indicates cross-β conformation. Proc Natl Acad Sci USA 83(2):503–507

    Article  PubMed  CAS  Google Scholar 

  15. Braak H, Braak E, Grundkeiqbal I, Iqbal K (1986) Occurrence of neuropil threads in the senile human-brain and in Alzheimers-disease—a 3 rd location of paired helical filaments outside of neurofibrillary tangles and neuritic plaques. Neurosci Lett 65(3):351–355

    Article  PubMed  CAS  Google Scholar 

  16. Reusche E, Ogomori K, Diabold J, Johannisson R (1992) Electron-microscopic study of paired helical filaments and cerebral amyloid using a novel en-bloc silver staining method. Virchows Arch, A Pathol Anat Histopathol 420(6):519–525

    Article  CAS  Google Scholar 

  17. Ferrari A, Hoerndli F, Baechi T, Nitsch RM, Götz J (2003) Beta-amyloid induces paired helical filament-like tau filaments in tissue culture. J Biol Chem 278(41):40162–40168

    Article  PubMed  CAS  Google Scholar 

  18. Crowther RA (1991) Straight and paired helical filaments in Alzheimer-disease have a common structural unit. Proc Natl Acad Sci USA 88(6):2288–2292

    Article  PubMed  CAS  Google Scholar 

  19. Inoue S, Kuroiwa M, Kisilevsky R (2002) AA protein in experimental murine AA amyloid fibrils: a high resolution ultrastructural and immunohistochemical study comparing aldehydfixes and cryofixed tissues. Amyloid 9(2):115–125

    PubMed  CAS  Google Scholar 

  20. Makovitzky J, Richter S (2009) The relevance of the aldehyde bisulfite toluidine blue reaction and its variants in the submicroscopic carbohydrate research. Acta Histochem 111(4):273–291

    PubMed  Google Scholar 

  21. Smith JF, Knowles TPJ, Dobson CM, MacPhee CE, Welland ME (2006) Characterization of the nanoscale properties of individual amyloid fibrils. Proc Natl Acad Sci USA 103(43):15806–15811

    Article  PubMed  CAS  Google Scholar 

  22. Dong M, Hovgaard MB, Mamdouh W, Xu S, Otzen DE, Besenbacher F (2008) AFM-based force spectroscopy measurements of mature amyloid fibrils of the peptide glucagons. Nanotechnology 19(38), article number: 384013

    Google Scholar 

  23. Porter AE, Knowles TPJ, Muller K, Meehan S, McGuire E, Skepper J, Welland ME, Dobson CM (2009) Imaging amyloid fibrils within cells using a Se-labelling strategy. J Mol Biol 392(4):868–871

    Article  PubMed  CAS  Google Scholar 

  24. Ferguson N, Becker J, Tidow H, Tremmel S, Sharpe TD, Krause G, Flinders J, Petrovich M, Berriman J, Oschkinat H, Fersht AR (2006) General structural motifs of amyloid protofilaments. Proc Natl Acad Sci USA 103(44):16248–16253

    Article  PubMed  CAS  Google Scholar 

  25. Iwata K, Fujiwara T, Matsuki Y, Akutsu H, Takahashi S, Naiki H, Goto Y (2006) 3D structure of amyloid protofilaments of beta2-microglobulin fragment probed by solid-state NMR. Proc Natl Acad Sci USA 103(48):18119–18124

    Article  PubMed  CAS  Google Scholar 

  26. Celej MS, Jares-Erijman EA, Jovin TM (2008) Fluorescent N-arylaminonaphthalene sulfonate probes for amyloid aggregation of alpha-synuclein. Biophys J 94(12):4867–4879

    Article  PubMed  CAS  Google Scholar 

  27. Cathcart ES, Shirahama T, Cohen AS (1967) Isolation and identification of a plasma component of amyloid. Biochim Biophys Acta 147(2):392–393

    CAS  Google Scholar 

  28. Romhanyi G, Deak G, Fisher J (1975) Aldehyde bisulfite-toluidine blue (ABT) staining as a topo-optical reaction for demonstration of linear order of vicinal OH groups in biological structures. Histochemistry 43(4):333–348

    Article  PubMed  CAS  Google Scholar 

  29. Makovitzky J, Richter S, Appel TR (2006) Topooptical investigations and enzymatic digestions on tissue-isolated amyloid fibrils. Acta Histochem 108(3):193–196

    Article  PubMed  CAS  Google Scholar 

  30. Jin L-W, Claborn KA, Kurimoto M, Geday MA, Maezawa I, Sohraby F, Estrada M, Kaminksy W, Kahr B (2003) Imaging linear birefringence and dichroism in cerebral amyloid pathologies. Proc Natl Acad Sci USA 100(26):15294–15298

    Article  PubMed  CAS  Google Scholar 

  31. Finzi L, Bustamante C, Garab G, Juang CB (1989) Direct observation of large chiral domains in chloroplast thylakoid membranes by differential polarization microscopy. Proc Natl Acad Sci USA 86(22):8748–8752

    Article  PubMed  CAS  Google Scholar 

  32. Garab G, Pomozi I, Jörgens R, Weiss G (2005) Method and apparatus for determining the polarization properties of light emitted, reflected or transmitted by a material using a laser scanning microscope. US Patent 6(856):391

    Google Scholar 

  33. Steinbach G, Pomozi I, Zsiros O, Menczel L, Garab G (2009) Imaging anisotropy using differential polarization laser scanning confocal microscopy. Acta Histochem 111(4):317–326

    Article  CAS  Google Scholar 

  34. Steinbach G, Pomozi I, Zsiros O, Pay A, Horváth GV, Garab G (2008) Imaging fluorescence detected linear dichroism of plant cell walls in laser scanning confocal microscope. Cytom A 73A(3):202–208

    Article  Google Scholar 

  35. Steinbach G, Pomozi I, Garab G, Makovitzky J (2006) Periodic twisted structure of amyloid fibrils revealed by differential polarization laser scanning microscopy. FEBS J 273(suppl 1):65

    Google Scholar 

  36. Pras M, Schubert M, Zucker-Franklin D, Rimon A, Franklin EC (1968) The characterization of soluble amyloid prepared in water. J Clin Invest 47(4):924–933

    Article  PubMed  CAS  Google Scholar 

  37. Linke RP (1983) Senile cardiac amyloid: biochemical and immunohistochemical results. In: Platt D (ed) Cardiology and ageing. Ist International Erlangen-Nürnberg Symposium on Experimental Gerontology 20–23 October 1982. Schattauer Verlag, Stuttgart, pp 81–97

    Google Scholar 

  38. Scherzinger E, Lurz R, Turmaine M, Mangiarini L, Hollenbach B, Hasenbank R, Bates GP, Davies SW, Lehrach H, Wanker EE (1997) Huntington-encoded polyglutamine expansions form amyloid-like protein aggregates in vitro and in vivo. Cell 90(3):549–558

    Article  PubMed  CAS  Google Scholar 

  39. Appel TR, Richter S, Linke RP, Makovitzky J (2005) Histochemical and topo-optical investigations on tissue-isolated and in vitro amyloid fibril. Amyloid 12(3):174–183

    Article  PubMed  CAS  Google Scholar 

  40. Romhányi G (1979) Selektive Darstellung sowie methodologische Möglichkeiten der Analyse ultrastruktureller Unterschiede von Amyloidablagerungen. Zentralbl Allg Pathol 123:9–16

    PubMed  Google Scholar 

  41. Romhanyi G (1958) Submicroscopic structure of elastic fibres as observed in the polarization microscope. Nature 182(4640):929–930

    Article  PubMed  CAS  Google Scholar 

  42. Garab G, van Amerongen H (2009) Linear and circular dichroism in photosynthesis research. Photosynth Res 101(2–3):135–146

    Article  PubMed  CAS  Google Scholar 

  43. Khurana R, Uversky VN, Nielsen L, Fink AL (2001) Is Congo Red an amyloid-specific dye? J Biol Chem 276(25):22715–22721

    Article  PubMed  CAS  Google Scholar 

  44. Szito T, Garab G, Mustárdy LA, Kiss JG, Faludi-Dániel A (1984) Increasing fluctuation in orientation of pigment protein complexes within photosynthetic membranes treated with linolenic acid. Photobiochem Photobiophys 8(4):239–249

    CAS  Google Scholar 

  45. Tinoco IJ, Mickols W, Maestre MF, Bustamante C (1987) Absorption, scattering, and imaging of biomolecular structures with polarized-light. Ann Rev Biophys 16:319–349

    Article  CAS  Google Scholar 

  46. Keller D, Bustamante C (1986) Theory of the interaction of light with large inhomogeneous molecular aggregates. II. Psi-type circular dichroism. J Chem Phys 84:2972–2979

    Article  CAS  Google Scholar 

  47. Chirico G, Langowski J (1996) Brownian dynamics simulations of supercoiled DNA with bent sequences. Biophys J 71(2):955–971

    Article  PubMed  CAS  Google Scholar 

  48. Bishop TC (2008) Geometry of the nucleosomal DNA superhelix. Biophys J 95(3):1007–1017

    Article  PubMed  CAS  Google Scholar 

  49. Bondarenko VA, Jiang YI, Studitsky VM (2003) Rationally designed insulator-like elements can block enhancer action in vitro. EMBO J 22:4728–4737

    Article  PubMed  CAS  Google Scholar 

  50. Kaminsky W, Jin L-W, Powell S, Maezawa I, Clabon K, Branham C, Kahr B (2006) Polarimetric imaging of amyloid. Micron 37(4):324–338

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Biological Research Center, Hungarian Academy of Sciences, P.O. Box 521, Szeged, 6701, Hungary

    Gábor Steinbach, István Pomozi, Dávid Péter Jánosa & Győző Garab

  2. Pi Vision Bt., Budapest, Hungary

    István Pomozi

  3. Department of Neuropathology, University of Heidelberg, Heidelberg, Germany

    Josef Makovitzky

  4. Medicopt Bt., Szeged, Hungary

    Győző Garab

Authors
  1. Gábor Steinbach
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. István Pomozi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Dávid Péter Jánosa
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Josef Makovitzky
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Győző Garab
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Győző Garab.

Electronic supplementary material

Below is the link to the electronic supplementary material

Supplemental Movie 1

Animated 3D image of the anisotropic architecture of isolated human amyloid sample (shown in Figure 1), reconstructed from fluorescence detected linear dichroism (FDLD) images of a series of optical sections with Z-spacings of 1 µm. Tilting, between -30° and +30°. (avi 31.5 mb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Steinbach, G., Pomozi, I., Jánosa, D.P. et al. Confocal Fluorescence Detected Linear Dichroism Imaging of Isolated Human Amyloid Fibrils. Role of Supercoiling. J Fluoresc 21, 983–989 (2011). https://doi.org/10.1007/s10895-010-0684-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10895-010-0684-3

Keywords

Search

Navigation