Neurodegenerative diseases such as Alzheimer’s and Parkinson’s are often accompanied by amyloid proteins in the brain that have become clumped or misfolded. At Washington University in St. Louis, a newly developed technique that measures the orientation of single molecules is enabling optical microscopy to reveal nanoscale details about the structures of these problematic proteins.
Research from the lab of Matthew Lew, assistant professor in the Preston M. Green Department of Electrical & Systems Engineering at the McKelvey School of Engineering, describing this new approach was published in Optica, The Optical Society’s journal for high impact research.
Biological and chemical processes are driven by complicated movements and interactions between molecules. Although most amyloid proteins may be non-toxic, the misfolding of even a few could eventually kill many neurons.
Lew’s lab has developed several microscopy methods that measure the orientation and location of fluorescent molecules attached to single proteins. The orientation information is obtained by measuring not only the location of fluorescence in the sample but also characteristics of that light, such as polarization, which are typically ignored in most other microscopy approaches.
The researchers used the powerful microscopy approach to measure the orientations of single fluorescent molecules bound to amyloid aggregates. Because there is no artificial linker between the fluorescent probes and amyloid surfaces, the probes’ binding orientation to the amyloid surfaces conveys information about how the amyloid protein itself is organized.
The researchers quantified how the orientations of each fluorescent molecule varied each time one attached to an amyloid protein. Differences in these binding behaviors can be attributed to structure differences between amyloid aggregates. Because the method provides single-molecule information, the researchers could observe nanoscale differences between amyloid structures without averaging out details of local features.
The researchers note that the set-up they used for orientation-localization microscopy consisted of commercially available parts that are accessible to anyone performing single-molecule super-resolution microscopy. Their analysis code is available online at: https://github.com/Lew-Lab/RoSE-O.
