Acceleration of nucleation of prion protein during continuous ultrasonication.

Although pulsatile irradiation of ultrasonication is frequently used for generating amyloid fibrils in vitro, the potential for inducing amyloid fibrillation of proteins during continuous ultrasonication is unknown. In this study, we implemented a continuous irradiation system and measured far-ultraviolet circular dichroism in a real-time manner. During the continuous ultrasonication, the conformation of full-length mouse prion protein (mPrP) was rapidly altered without a lag time and electron microscopy revealed that distorted fibrils, ß-oligomers and amorphous aggregates were formed at pH 2.2, 4.0 and 9.1, respectively. Similarly, hen egg white lysozyme formed distorted fibrils and small and large amorphous aggregates at pH 2.2 and 7.1 and 11.9, respectively, without a lag time. The concentration dependencies of the initial rates were different between the two systems. The aggregate formation of mPrP followed a first-order reaction, whereas that of lysozyme followed the zeroth-order reaction. Importantly, the reactions were immediately stopped by switching off ultrasonication, and restarted instantaneously when ultrasonication was restarted. Thus, the continuous ultrasonication significantly accelerates the nucleations of mPrP and lysozyme aggregates by the interaction between monomer and cavitation bubble. These cavitation bubbles may act as catalysts that decrease the activation free energy for nucleation, which is low in mPrP and high in lysozyme.