Amyloid fibril formation by a partially structured intermediate state of alpha-chymotrypsin.

Here we investigated the effects of 2,2,2-trifluoroethanol (TFE) on the structure of alpha-chymotrypsin. The protein aggregates maximally in 35% (v/v) TFE. Congo red and thioflavin-T binding experiments suggest that the aggregates induced by TFE have amyloid-like properties, and transmission electron microscopy data show that these aggregates have a fibrilar morphology. Fluorescence, circular dichroism, anilino-8-napthalene sulfonate binding, and Fourier-transformed infrared spectroscopy data suggest that formation of a partially structured intermediate state precedes the onset of the aggregation process. The native beta-barrel structure of alpha-chymotrypsin appears to be disrupted in the partially structured intermediate state in favour of a non-native extended beta-sheet conformation with exposed hydrophobic surfaces. The protein becomes "sticky" under these conditions and aggregates into amyloid-like structures. The data support the hypothesis that amyloid formation involves the ordered self-assembly of partially folded species that are critical soluble precursors of fibrilar aggregates.