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On the Protein Fibrillation Pathway: Oligomer Intermediates Detection Using ATR-FTIR Spectroscopy.
- Source :
-
Molecules (Basel, Switzerland) [Molecules] 2021 Feb 12; Vol. 26 (4). Date of Electronic Publication: 2021 Feb 12. - Publication Year :
- 2021
-
Abstract
- Oligomeric intermediates on the pathway of amyloid fibrillation are suspected as the main cytotoxins responsible for amyloid-related pathogenicity. As they appear to be a part of the lag phase of amyloid fibrillation when analyzed using standard methods such as Thioflavin T (ThT) fluorescence, a more sensitive method is needed for their detection. Here we apply Fourier transform infrared spectroscopy (FTIR) in attenuated total reflectance (ATR) mode for fast and cheap analysis of destabilized hen-egg-white lysozyme solution and detection of oligomer intermediates of amyloid fibrillation. Standard methods of protein aggregation analysis- Thioflavin T (ThT) fluorescence, atomic force microscopy (AFM), and 8-anilinonaphthalene-1-sulphonic acid (ANS) fluorescence were applied and compared to FTIR spectroscopy data. Results show the great potential of FTIR for both, qualitative and quantitative monitoring of oligomer formation based on the secondary structure changes. While oligomer intermediates do not induce significant changes in ThT fluorescence, their secondary structure changes were very prominent. Normalization of specific Amide I region peak intensities by using Amide II peak intensity as an internal standard provides an opportunity to use FTIR spectroscopy for both qualitative and quantitative analysis of biological samples and detection of potentially toxic oligomers, as well as for screening of efficiency of fibrillation procedures.
- Subjects :
- Amyloid ultrastructure
Amyloidogenic Proteins ultrastructure
Animals
Chickens
Fluorescence
Microscopy, Atomic Force
Muramidase ultrastructure
Protein Structure, Secondary
Spectroscopy, Fourier Transform Infrared
Amyloid chemistry
Amyloidogenic Proteins chemistry
Benzothiazoles chemistry
Muramidase chemistry
Subjects
Details
- Language :
- English
- ISSN :
- 1420-3049
- Volume :
- 26
- Issue :
- 4
- Database :
- MEDLINE
- Journal :
- Molecules (Basel, Switzerland)
- Publication Type :
- Academic Journal
- Accession number :
- 33673072
- Full Text :
- https://doi.org/10.3390/molecules26040970