Conformational bifurcation of tau R3-R4 oligomers underlies fibril polymorphism.

Tau aggregation into amyloid fibrils is a central pathological feature of multiple neurodegenerative diseases, yet identical tau sequences can adopt structurally distinct fibril conformations associated with different disorders. How such disease-specific folds emerge from the same sequence remains poorly understood. Here, we use metadynamics and all-atom molecular dynamics simulations to elucidate the folding mechanisms of tau trimers comprising the R3-R4 region, the minimal aggregation nucleus of tau fibrils. By constructing the folding free-energy landscape within a pre-organized fibril-like scaffold, we identify a shared, partially folded intermediate from which two energetically comparable minimum free-energy pathways diverge, leading to either a compact Alzheimer's disease-like or a more open chronic traumatic encephalopathy-like C-shaped conformation. Structural analysis reveals that the β4-β6 triangular region acts as the central determinant of this bifurcation. Hydrophobic-core rearrangements within this region provide the primary energetic driving force for compaction, whereas the conformational flexibility of the 332PGGG335 hinge modulates whether this tendency can be structurally realized. Additional simulations show that stable formation of the triangular region requires cooperative multichain interactions; reducing oligomer size destabilizes this scaffold and abolishes ordered folding. Together, our results establish a mechanistic framework in which identical tau sequences access alternative folding routes through a common intermediate, explaining the emergence of disease-specific fibril polymorphism at the oligomeric level and highlighting early folding intermediates as potential targets for therapeutic intervention.