Determination of α-Synuclein Protein Interactions by μMap Photo-proximity Labeling

Fibrillar aggregates of the natively disordered protein -synuclein (S) are hallmarks of Parkinsons disease and related neurodegenerative disorders termed synucleinopathies. Here, we used micromap ({micro}Map) photo-proximity labeling to determine the interactomes of S monomers and fibrils in mouse brain lysate to better understand both the loss of healthy function and gain of toxic function aspects of synucleinopathies. Several S variants were synthesized and characterized, showing that the small size (1 kDa) of the Ir catalyst attached through a Cys-maleimide linkage makes it minimally-perturbing to S, with a narrow labeling radius that allows one to identify interactome differences between different regions of S. Monomer and fibril interactomes were compared to each other and to previous proximity labeling data sets for validation and several examples of further investigations are demonstrated, including Western blotting, super-resolution microscopy, and {micro}Map in primary neurons.