Ultrafast and Nonlinear Spectroscopy Utilized as a Sensitive Probe for Amyloid Peptide Aggregation.

Abstract

The development of new sensitive methods for the detailed collection of structural and conformational information about amyloid peptides is critical to elucidating the fundamental aggregation processes that lead to neurodegenerative disease. This research project demonstrates the development of a combined methodology utilizing nonlinear and ultrafast time-resolved spectroscopies for the study of aggregating peptides. Synthetic organic chemistry was crucially implemented to design a biologically compatible chromophore similar in structure to that found in green fluorescent protein (GFP) but one possessing superior two-photon absorption (TPA) characteristics. The photophysical advantages of tailoring a chromophore to address the challenges of studying a dynamic system like amyloid-beta were clearly evident in the strong environmental sensitivity of the chromophore for the changing conformations of amyloid-beta. The overall impact in the work lies with the significant two-photon absorption enhancement observed with labeled amyloid-beta 1-42 that was found to correspond directly to structural features of the aggregating peptides. This sensitivity demonstrates some of the advantages of a two-photon methodology over a traditionally used method like circular dichroism. A two-photon technique has superior low-concentration sensitivity and no limitations due to background sources of absorption. The complete two-photon absorption and time-resolved fluorescence methodology permits detailed conformational and peptide aggregate characterization of the evolution from early oligomer to fibril formation, something that could have a huge impact toward therapeutic intervention or fundamental understanding of Alzheimer’s disease. As a result of this work, a firm foundation for the continued development of nonlinear and time-resolved ultrafast spectroscopies in the investigation of aggregating peptide systems has been established with the known capability to perform fundamental investigations of amyloid systems that are difficult with other biochemical techniques. Many direct applications for in vivo imaging may be envisioned using two-photon-based methodologies including cancer targeted monitoring as well as peptide aggregation associated with Alzheimer’s and Parkinson’s disease. Two-photon imaging has the capability to permit detailed tracking of the evolution of disease beginning with some initial misfolded monomer to the formation of toxic oligomers and later fibril structure. This could have a profound impact on the ability to detect and prevent the development of neurodegenerative disease.