Picosecond dynamics of polymorphs of lysozyme aggregates with different levels of cytotoxicity

Lysozyme amyloidosis is a hereditary severe disease, where lysozyme amyloid deposits in liver and kidney cause massive hemorrhage, resulting in the death of a patient. Amyloid fibrils are self-assembled protein filaments with core regions rich in β-sheet. It has been shown that lysozyme fibrils show structural polymorphism depending on fibrillation conditions and each polymorph shows different levels of cytotoxicity: fibrils formed at neutral pH are more cytotoxic than those formed at acidic pH. Cytotoxicity originates from interactions between fibrils and cell membranes. In particular, it is the amino acid side-chains of the fibrils that directly interact with the components of the membranes. It is thus important to characterize the mobility of side-chains of lysozyme polymorphs to gain insights into the molecular mechanism of cytotoxicity.In this study, we focused on the polymorphism of hen egg white lysozyme (HEWL), a model for studying lysozyme amyloidosis in humans and prepared D2O hydrated powder samples of two HEWL amyloid polymorphs formed at pH6.0 (hereafter called Fib6) or 2.7 (Fib2), which are known to show higher and lower levels of cytotoxicity, respectively. We carried out elastic incoherent neutron scattering (EINS) measurements on these samples using the IN13 spectrometer at Institut Laue-Langevin in France in the temperature range from 20 K to 310 K. In the first analysis, temperature dependences of the mean square displacements (MSDs) of atomic motions in the proteins were evaluated from the EINS spectra. There were, however, no significant differences in the extracted MSD values between the two samples in the temperature range studied albeit the difference in the shape of the EINS curves. Next, analysis based on the mean square positional fluctuations (MSPFs) was conducted, which uses all the data points in the EINS curves and thus provides more detailed dynamical information than the MSD analysis. It was found that whereas the MSPF values of atomic motions are similar between the two samples, each polymorph shows a different degree of motionalheterogeneity: Fib6 showed a larger fraction of atoms undergoing motions with larger amplitudes than Fib2. Furthermore, application of the Bicout -Zaccai model showed that ΔG of atomic motions tends to be lower for Fib6, suggesting that the energy barrier of atomic motions is lower for Fib6. These results suggest that the differences in molecular dynamics between lysozyme polymorphs lie in the distribution of local atomic motions and these dynamical differences would modulate the way by which the polymorphs interact with cell membranes, contributing to the differences in the level of cytotoxicity.Journées de la Diffusion Neutronique 202