Surface characterization of amyloid fibrils
The formation of amyloid fibrils is due to the misfolding of proteins and is discussed in the context of the development of neurodegenerative diseases. A detailed characterization of such protein aggregates with tip-enhanced Raman spectroscopy (TERS) can provide an important contribution to the understanding of the chemical composition on the nanometer scale.
By: TANJA DECKERT-GAUDIG, VOLKER DECKERT
Amlyoid fibrils are protein aggregates whose deposits can irreversibly damage cells and tissues and are discussed as triggers of serious diseases such as type II diabetes or Alzheimer’s disease. The fibrous twisted structures have a length of several 100 nm to several micrometers. So far, neither the exact formation mechanism nor the trigger of fibrillation are known. Accurate chemical characterization of fibril surfaces is of great interest in this context, since information about this interface can provide clues to fibril formation and this is where the first contact with the environment occurs. In recent years, tip-enhanced Raman spectroscopy (TERS) has proven to be a suitable analytical method in this field. Scanning of the fibril surface with an atomic force microscope and simultaneous detection of Raman spectra allow morphological and chemical characterization in one experiment – on a single-fibril basis with nanometer spatial resolution. The intrinsically high specificity of Raman spectroscopy enables the classification and discrimination of amino acids and secondary structures on fibrils. For example, on amyloid fibrils of insulin and amylin, it was demonstrated that the surfaces always contain a mixture of α-helix/disordered structures and β-sheet structures. These results extend the knowledge of amyloid fibrils whose core is composed exclusively of β-sheet structures.
In TERS experiments on amylin fibrils generated at different pH values, it was shown that the acidity of the environment has an effect on the surface chemical composition of the fibrils. Thus, fibrils at pH 2 exhibit a lower proportion of α-helix/disordered structures than structures grown at pH 7.8. The presence of mixed structures on the fibril surface is consistent with theoretical models and could not be visualized by any other method so far.
At the same time, the high sensitivity of TERS and the precision of tip positioning on a fibril allows the detection of secondary structure transitions, as shown in the figure. Here, spectra were systematically recorded along a profile line on an amylin fibril at 0.5 nm intervals. Based on the location of the amide I band characteristic of a peptide bond, a change from ordered β-sheet structures at the beginning of the measurement to α-helix/disordered structures can be seen. The detection of molecular structural changes within a few nanometers is only possible due to the high spatial spatial resolution capability of TERS. This feature, together with the aforementioned specificity and sensitivity, allow direct surface chemical analysis of a wide variety of biological, organic, and inorganic samples, which is not as possible with other methods.
In a recent study, the fibrillation of amylin was investigated in the presence of a lipid double membrane. Interactions of this peptide with lipids of the cell membrane are thought to play a special role in the development of type II diabetes. As shown by the experiments, lipids could indeed be detected on single fibrils based on their marker bands in the TERS spectra. These results are a promising starting point for future experiments to study amyloid fibrils on cells and in the presence of potential reagents to inhibit fibril formation.
Funded by: EU, COST