We study the fibrillation pathway of ovalbumin protein and report the simultaneous formation of several types of fibrils, with clear structural and physical differences. We compare the fibrillation mechanisms at low pH with and without salt, and follow the kinetics of fibrils growth by atomic force microscopy (AFM), static and dynamic light scattering (SLS, DLS), and small-angle X-ray scattering (SAXS). We show that, among the morphologies identified, long semiflexible amyloid fibrils (type I), with persistence length Lp∼3 μm, Young's modulus E∼2.8 GPa, and cross-β structure are formed. We also observe much more flexible fibrils (type III, Lp∼63 nm), that can assemble into multistranded ribbons with time. They show significantly lower intrinsic stiffness (1.1 GPa) and a secondary structure, which is not characteristic of the well-ordered amyloids, as determined by circular dichroism (CD), wide-angle X-ray scattering (WAXS), and attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR). In between these two main classes of fibrils, a third family, with intermediate flexibility (type II, Lp∼300 nm), is also resolved.

Special issue in Honour of Pierre TURQInternational audienceWe used several complementary experimental and theoretical tools to characterise the charge properties of well-definedmaghemite nanoparticles in solution as a function of the volume fraction. The radius of the nanoparticles is equal to 6 nm.The structural charge was measured from chemical titration and was found high enough to expect some counterions tobe electrostatically attracted to the surface, decreasing the apparent charge of the nanoparticle. Direct-current conductivitymeasurements were interpreted by an analytical transport theory to deduce the value of this apparent charge, denoted here by‘dynamic effective charge’. This dynamic effective charge is found to decrease strongly with the volume fraction. In contrast,the ‘static’ effective charge, defined thanks to the Bjerrum criterion and computed from Monte Carlo simulations turns outto be almost independent of the volume fraction. In the range of Debye screening length and volume fraction investigatedhere, double layers around nanoparticles actually interact with each other. This strong interaction between nanocolloidalmaghemite particles is probably responsible for the experimental dependence of the electrokinetic properties with the volumefraction

Confocal laser scanning microscopy (CLSM) is the most popular technique for mapping the subcellular distribution of a fluorescent molecule and is widely used to investigate the penetration properties of exogenous macromolecules, such as cell-penetrating peptides (CPPs), within cells. Despite the membrane-association propensity of all these CPPs, the signal of the fluorescently labeled CPPs did not colocalize with the plasma membrane. We studied the origin of this fluorescence extinction and the overall consequence on the interpretation of intracellular localizations from CLSM pictures. We demonstrated that this discrepancy originated from fluorescence self-quenching. The fluorescence was unveiled by a “dilution” protocol, i.e. by varying the ratio fluorescent/non-fluorescent CPP. This strategy allowed us to rank with confidence the subcellular distribution of several CPPs, contributing to the elucidation of the penetration mechanism. More generally, this study proposes a broadly applicable and reliable method to study the subcellular distribution of any fluorescently labeled molecules.