Nanoscale inspection of toxic amyloids by tip-enhanced Raman spectroscopy

Plasmon-enhanced spectroscopies rely on the resonance effects between a laser excitation and free conduction electrons in noble metal nanostructures, permitting to enhance the optical responses of molecules in their close proximity. Surface-Enhanced Raman Spectroscopy (SERS) is an ultrasensitive analytical technique that couples the unique features of Raman Spectroscopy in providing a description of chemical composition and structure of molecules with a huge signal enhancement due to isolated or assembled silver or gold nanoparticles. As a result, identification of trace amounts of molecules, including those of biomedical interest, becomes feasible [1-5]. The intrinsic chemical specificity of Raman Spectroscopy and the high signal sensitivity of SERS can be combined with the nanoscale spatial resolution of scanning probe microscopy (SPM) giving rise to Tip-Enhanced Raman Spectroscopy (TERS). In TERS the huge electromagnetic field localized on the apex of a sharp metallized tip is exploited to achieve compositional and structural information from the surface of nanosized samples [6]. In this work we show how to benefit from TERS as a surface-sensitive tool with spatial resolution on the nanoscale to inspect the spatial organization and surface character of individual protein oligomers causing cellular dysfunction and neurotoxicity. Aggregation processes of proteins leading to the formation of amyloid oligomers are nowadays considered primarily responsible for promoting synaptic failure and neuronal death associated with neurological disorders such as Alzheimer's and Parkinson's diseases. Therefore, unravelling the relationship between structure and neurotoxicity of protein oligomers becomes of primary importance in understanding the pathogenesis of the disease as well as in developing novel diagnostic and therapeutic strategies toward the earliest and pre-symptomatic stages of the disease. Our TERS investigation provides strong evidence of the presence of characteristic chemostructural determinants in toxic oligomers, which sheds new light on the mechanism by which they cause cellular impairment. Specifically, we provide direct assignment of specific aminoacid residues that are exposed on the surface of toxic amyloids, while appear buried in nontoxic amyloid forms. These residues, thanks to their outward disposition, might represent structural factors driving the pathogenic behaviour, including affecting cell membrane integrity and specific signal pathways in neurodegenerative conditions.