Characterization of intermediate states in the fibrillogenesis of odorant-binding proteins.

The transition of β-barrel proteins from a soluble to an amyloid form is biologically significant in some cases but may lead to functional activity loss. In particular, odorant-binding proteins' (OBPs) fibrils are unable to bind odorant molecules potentially contributing to olfactory dysfunction. As shown previously, OBPs' fibrillogenesis is initiated by uncoupling of protein C-terminal fragment from the β-barrel and exposing amyloidogenic sites. However, further structural transformations of OBPs are not fully understood. Here we first identified two intermediate aggregated states of OBPs: one formed by intact β-barrels, nontoxic to mammalian cells and easily dissociated by boiling and detergents; the other is prefibrillar, formed by degraded β-barrels with restructured β-strands, having almost comparable cytotoxicity but lower stability compared to mature amyloids. Obtained results revealed the β-barrel "opening" and alignment in register of its β-strands early in aggregation. Comparing the aggregation processes of bovine OBP and its mutant variant differing in the C-terminal fragment mobility showed the influence of its dynamics/orientation on the conjugation of amyloidogenic regions triggering fibril formation. The characterized properties and formation mechanisms of intermediate states in amyloidogenesis of β-barrel proteins are relevant for finding ways to prevent pathological aggregation and identifying ways to regulate the physiological fibrils assembly/disassembly.