Modulation of Surface-Catalyzed Secondary Nucleation during Amyloid Fibrillation of Hen Egg White Lysozyme by Two Common Surfactants.

Amyloid fibrillation by hen egg white lysozyme (HEWL) under the influences of two common surfactants, sodium dodecyl sulfate (SDS) and Triton X-100 (TX-100), was investigated with atomic force microscopy (AFM) and Fourier transform infrared (FTIR) spectroscopy. Detailed AFM investigations indicated that both SDS and TX-100 were able to induce some significant morphological changes of HEWL amyloid fibrils. Both SDS and TX-100 could induce a morphological change which was characterized with alternating fibril regions with increased thicknesses along the fibril axis. In addition, TX-100 was also able to induce a morphological change which was characterized with fibril branching. In contrast, the positively charged cetyltrimethylammonium bromide displayed no such effect as compared with the negatively charged SDS and the nonionic TX-100. These intriguing modulation effects of SDS and TX-100 on amyloid fibrillation were hypothesized to be due to surfactant-induced surface-catalyzed secondary nucleation owing to the noncovalent interaction between the surfactants and HEWL. The proposed hypothesis was further supported by FTIR spectroscopic investigation, which demonstrated the formation of a SDS-amyloid fibril complex and TX-100-amyloid fibril complex. Detailed FTIR analysis suggested that the tail group of SDS associated with amyloid fibrils is more disordered similar to that of SDS in aqueous solution, while the headgroup of SDS appears to interact with amyloid fibrils strongly similar to that of SDS in the solid state, and the ether groups of TX-100 associated with amyloid fibrils experienced a different microenvironment as compared with that of neat TX-100. Additional control experiments with FTIR spectroscopy revealed that the interactions of SDS/TX-100 with HEWL amyloid fibrils were different from those of SDS/TX-100 with HEWL amorphous aggregates. In addition, the ß-sheet structures of the HEWL amyloid fibrils were found to be increased due to the influences of SDS and TX-100. Our work provides new insight into the modulation effects of surfactants on amyloid fibrillation.

A structural model of the hierarchical assembly of an amyloid nanosheet by an infrared probe technique.

Rational design of amyloid-based materials requires structural insight into such materials. Here, we explore the use of a side-chain-based infrared (IR) probe technique combined with atomic force microscopy, Raman spectroscopy, UV-Vis spectroscopy, and thermogravimetric analysis coupled with mass spectrometry to elucidate the structural details of an amyloid nanosheet formed by an Aß(16-22) variant, KLVXFAK, where X is p-cyanophenylalanine with its side-chain cyano group being an IR probe. Through the structural constraints obtained with the combined tools, we are able to propose a novel structural model for the amyloid nanosheet. The nanosheet can be viewed as a stack of class 7-type steric-zipper-like amyloid structures with a unique sheared intersheet arrangement: the ß-sheets are stacked along a zippering axis with each individual ß-sheet sheared relative to its adjacent ß-sheets by two residues through the cyano-lysine intersheet hydrogen bond. With such a configuration, the side view of the nanosheet is similar to that of a stack of tilted-aligned roof tiles with each individual ß-sheet being each tile. In addition, this work provides a nice example of how to utilize the side-chain-based IR probe technique combined with other supplemental tools to build a hierarchical structural model for a complex amyloid assembly.

Kinetic Mechanism of Thioflavin T Binding onto the Amyloid Fibril of Hen Egg White Lysozyme.

Thioflavin T (ThT) is widely used as a fluorescent probe for amyloid fibril detection. Yet the exact kinetic mechanism of ThT binding onto amyloid fibril remains elusive. Previously reported kinetic studies using ThT-fluorescence-detected kinetic design suggested two completely different ThT-binding mechanisms. In one study, a multistep sequential binding mechanism onto a single ThT-binding site was suggested. In another study, a one-step parallel binding mechanism onto multiple ThT-binding sites was suggested. The discrepancy is likely due to the incapability of ThT-fluorescence-detected kinetic design to differentiate the two above-mentioned mechanisms. Considering the weakness of the ThT-fluorescence-detected approach, we investigated the ThT-binding mechanism onto the amyloid fibril of hen egg white lysozyme (HEWL) using a new approach, ThT-absorbance-detected kinetic design. Our new results suggest that ThT binds to HEWL fibril through the one-step parallel binding mechanism. We hope our work can offer some new insights into the interactions between dye molecules and amyloid fibrils.