Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Fluorescent fusion proteins as probes to characterize tau fibril polymorphism
Linköping University, Department of Physics, Chemistry and Biology, Chemistry. (Hammarström Lab)
2019 (English)Independent thesis Advanced level (degree of Master (Two Years)), 40 credits / 60 HE creditsStudent thesis
Abstract [en]

Alzheimer's disease (AD) is a large and growing problem and while we today lack a full understanding of this disease, we know that the protein tau and the amyloid fibrils it forms play a central role in its development. We also know that these fibrils can have different morphologies in different diseases and that fibrils produced in vitro not necessarily adopt any of the morphologies found in patients. This means there is a need for more pathologically relevant fibrils in vitro to be able to understand this disease better. One approach to satisfy this need is to use fibrils found in patients as seeds and thus transfer their morphology to recombinantly purified protein. To facilitate this process this study has attempted to develop a way to differentiate between different fibril morphologies using a FRET based system. This involves fluorescent fusion proteins (tau-EXFPs) and fluorescent amyloid probes as well as seeding experiments with pseudo wild type tau (PWT) and tau with the P301L mutation. Greater differences in terms of fibrillation rates and ThT fluorescence between PWT and P301L was shown than previously reported between WT and P301L. They were also shown to differ in fibril morphology in TEM. The ThT fluorescence intensity was to a certain degree transferable from PWT to P301L by seeding. Furthermore, this study confirms that the tau-EXFP fusion protein can be incorporated into amyloid fibrils and strongly suggests that a FRET effect between EXFP and BTD14 (as well as X34 and ThT) can be achieved. It also demonstrates differences in FRET efficiency between PWT and P301L fibrils using FLIM. These results indicate that a FRET based approach could be a useful method to discern different fibril morphologies from each other, but further measurements and optimization are needed before this method could be reliably applied. The fusion proteins could also be used to investigate tau spreading in vivo, e.g. in D. melanogaster. To find suitable FRET partners to the fusion proteins, a ligand screen was conducted. This could be used as an alternative to the FRET method. With the right selection of fluorescent amyloid probes, a unique fingerprint for each fibril morphology could maybe be generated and fulfill the same intended function as the FRET method.

Place, publisher, year, edition, pages
2019. , p. 76
Keywords [en]
Alzheimer’s disease (AD), amyloids, FRET, tau, MAPT, fibrillation kinetics, seeding, fusion protein, EGFP, ECFP, EYFP, ThT, X-34, BTD14
National Category
Structural Biology
Identifiers
URN: urn:nbn:se:liu:diva-158263ISRN: LITH-IFM-A-EX--19/3627--SEOAI: oai:DiVA.org:liu-158263DiVA, id: diva2:1331993
Subject / course
Chemistry
Supervisors
Examiners
Available from: 2019-06-28 Created: 2019-06-27 Last updated: 2019-06-28Bibliographically approved

Open Access in DiVA

fulltext(4812 kB)15 downloads
File information
File name FULLTEXT01.pdfFile size 4812 kBChecksum SHA-512
1be969b58e42ca6159bce2125e40c6a7b34b1d70ca66d5167719aa7c001773fdeeebbe5d3cc77fdbf7337c90f69fb154cb946be5f4da9c6c4c66d253662f2977
Type fulltextMimetype application/pdf

Search in DiVA

By author/editor
Lindberg, Max
By organisation
Chemistry
Structural Biology

Search outside of DiVA

GoogleGoogle Scholar
Total: 15 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

urn-nbn

Altmetric score

urn-nbn
Total: 205 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf