De novo Design of Polymorph-Specific Binders Targeting α-Synuclein Fibrils

Alpha-synuclein (aSyn) aggregation into amyloid fibrils is a hallmark of synucleinopathies, a group of neurodegenerative disorders that includes Parkinson’s disease. Beyond the central role of fibrils in disease pathology and propagation, distinct aSyn fibril polymorphs have been associated with different synucleinopathies, presenting opportunities for differential diagnosis and structure-based disease-specific phenotyping. Here, we present AmyBind, a computational pipeline for the de novo design of mini-protein binders that target polymorph-specific structural features of amyloid fibrils. Using this approach, we developed two aSyn fibril-specific binders, one of which exhibited polymorph-specificity. Neither binder interfered with fibril elongation in vitro , consistent with their designed binding mode on lateral fibril surfaces. In cell-based models, both binders showed polymorph-specific colocalization but did not affect fibril uptake or seeding, indicating their utility as fibril-selective labeling agents in cellular contexts. These findings demonstrate the feasibility of structure-guided, polymorph-specific amyloid targeting and provide a foundation for developing early-stage diagnostics and therapeutics for synucleinopathies and other amyloid-related diseases.