The folding and acquisition of native structure of proteins is critical to life-enabling biological processes. However, protein misfolding can lead to toxic amyloid aggregates, linked to the onset of neurodegenerative disorders such as Alzheimer’s disease. To shed light on the molecular basis of protein function and malfunction, it is crucial to access structural information of single protein assemblies and aggregates under native conditions. Conformation-sensitive spectroscopic methods lack the spatial resolution and sensitivity necessary for characterizing heterogeneous protein aggregates in water.<br/>Photothermal induced resonance (PTIR) is an emergent AFM technique that provides IR spectra and maps with nanoscale resolution, thus opening a nanoscale observation window on the conformation of peptides and proteins, but its implementation is water has been challenging.<br/>After a brief PTIR introduction to PTIR I will show that such limitations can be overcome and that, for the first time, PTIR can be leveraged to acquire conformationally-sensitive nanoscale infrared spectra in water with high signal to noise ratio (SNR &gt; 70). Using conventional AFM probes, we study supramolecular aggregates of diphenylalanine (FF), the core recognition module of the Alzheimer's disease β-amyloid peptide, and its derivative boc-diphenylalanine (Boc-FF). We demonstrate that nanoscale resolved IR spectra and maps in air, and water can be obtained with comparable SNR and lateral resolution, enabling accurate identification of the chemical and structural state of the aggregates at the single aggregate level. Our data confirm that the FF and Boc-FF fibrils differ structurally at the individual fibril level due to the additional presence of α-helical conformation in the Boc-FF with respect to the pure β-sheet conformation of FF.<br/>In the final part of my talk, I will introduce a novel nano-sized AFM probe that leverages integrated cavity opto-mechanics to improve, in air, the PTIR sensitivity (50x), time resolution (1500x) and throughput (&gt;2500x) concurrently. I will highlight the potential of these novel nanoscale probes to also dramatically improve the PTIR performance in liquid thanks to a greatly reduced cantilever drag.

Determination of polypeptides conformation in water with infrared nano-spectroscopy

2019 Fall National Meeting

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