Hydration water mobility is enhanced around tau amyloid fibers

The paired helical filaments (PHF) formed by the intrinsically disordered human protein tau are one of the pathological hallmarks of Alzheimer disease. PHF are fibers of amyloid nature that are composed of a rigid core and an unstructured fuzzy coat. The mechanisms of fiber formation, in particular the role that hydration water might play, remain poorly understood. We combined protein deuteration, neutron scattering, and all-atom molecular dynamics simulations to study the dynamics of hydration water at the surface of fibers formed by the full-length human protein htau40. In comparison with monomeric tau, hydration water on the surface of tau fibers is more mobile, as evidenced by an increased fraction of translationally diffusing water molecules, a higher diffusion coefficient, and increased mean-squared displacements in neutron scattering experiments. Fibers formed by the hexapeptide ³⁰⁶VQIVYK³¹¹ were taken as a model for the tau fiber core and studied by molecular dynamics simulations, revealing that hydration water dynamics around the core domain is significantly reduced after fiber formation. Thus, an increase in water dynamics around the fuzzy coat is proposed to be at the origin of the experimentally observed increase in hydration water dynamics around the entire tau fiber. The observed increase in hydration water dynamics is suggested to promote fiber formation through entropic effects. Detection of the enhanced hydration water mobility around tau fibers is conjectured to potentially contribute to the early diagnosis of Alzheimer patients by diffusion MRI.Amyloid fibers are the most stable forms of ordered protein aggregates. They have attracted much attention because of their implication in so-called conformational diseases, which include a variety of neurodegenerative disorders (1). Consequently, means of hindering or reversing fiber formation are actively researched (2). Pathological fibers are often formed by intrinsically disordered proteins (IDPs) that lack a well-defined 3D structure in their native state and are best described by an ensemble of different conformations (3). The human protein tau is an IDP that normally regulates microtubule stability in neurons. When tau aggregates, it forms paired helical filaments (PHF) that are one of the two histological hallmarks of Alzheimer disease (AD) (4, 5). As yet, and despite considerable effort over the past 30 y, the understanding of tau fibrillation in AD and other taupathies remains largely incomplete (6). The longest human tau isoform, htau40, is composed of 441 amino acid residues and is organized into several domains (see Fig. 1), including the repeat domains R1−R4 (residues 244–369) that constitute, together with the P1 and P2 domains, the microtubule binding regions (7). Essential for the nucleation of tau fibers is the presence of hexapeptides (²⁷⁵VQIINK²⁸⁰ and ³⁰⁶VQIVYK³¹¹) in R2 and R3 (8) that have a high propensity to form β-structures. Although precise structures of tau PHF remain unknown (6), they can be divided into two structurally different regions (see Fig. 1): (i) a rigid β-rich core (denoted as the fiber core domain), which is essentially composed of the four repeat domains, and (ii) the remainder, the so-called fuzzy coat, which is highly flexible (9–11).Open in a separate windowFig. 1.Schematic representation of the tau isoform htau40 in its monomeric (Top) and fibrillated (Bottom) forms. The microtubule binding domain is roughly composed of the four repeat domains R1−R4 (residues 244–369) and the proline-rich domains P1 and P2. R1−R4 constitute the core domain, which forms cross-β structures as well as steric zippers in the fiber, whereas the rest of the protein is referred to as the fuzzy coat domain, which remains disordered in the fiber form. The amyloidogenic hexapeptide ³⁰⁶VQIVYK³¹¹ can be used as a model for the fiber core.Water is known to play key roles in protein folding, stability, and activity (12). It mediates protein−protein and protein−DNA recognition, is involved in allostery, partakes in enzymatic reactions and proton and electron transfer, and more generally plasticizes biological macromolecules by providing their surface with an extensive and highly dynamic network of hydrogen bonds. Compared with folded proteins, tau has been shown to have a stronger coupling with its hydration water (13). However, very little is known about the role water plays in protein aggregation in general and in tau fibrillation in particular. A recent study on two different amyloid systems concluded that water plays a key role in fiber growth and polymorphism, inter alia through entropic effects (14). A study by Chong and Ham (15) highlighted the role of water in protein aggregation propensity by revealing a tight relation between the hydration free energy of a protein and its propensity to aggregate.Among the experimental methodologies available to study protein hydration water, neutron scattering (NS) stands out owing to its pronounced sensitivity to motions of hydrogen atoms. Indeed, hydrogen atoms incoherently scatter neutrons about two orders of magnitude more strongly than all other atoms present in a biological sample, including deuterium atoms. Consequently, NS has been widely used to study bulk and confined water at room temperature (16), hydration water of peptides (17), proteins (18–21), and water inside cells (22). More specifically, NS probes atomic motions on the nanosecond to picosecond timescales and on the angstrom length scales (23), thus ensuring the time and space resolution necessary for investigating water dynamics with atomistic detail. Elastic incoherent NS (EINS) reflects the global dynamics averaged over all atoms but does not provide any information on the nature of the observed motions. Quasi-elastic NS (QENS), however, allows the quantification of energy exchanges between the sample and the neutron beam and provides quantitative information about the nature of motions observed. Because of a pronounced isotope effect, the replacement of hydrogen by deuterium atoms effectively masks the labeled part of a sample in incoherent NS experiments. Perdeuteration of proteins (i.e., deuteration of the entire protein) hydrated in H₂O thus puts the focus on hydration water dynamics by minimizing the protein contribution to the NS signal. All-atom molecular dynamics (MD) simulation is a useful complement to NS because both methods probe atomic motions on the same time and length scales. Whereas incoherent NS provides an accurate measure of the average dynamics of hydrogen atoms throughout the sample, MD simulations provide atomic-scale insight into motions occurring within particular space and time windows of interest (24).Here we experimentally and computationally address the effect of tau fiber formation on the dynamics of its surrounding hydration water. We produced perdeuterated htau40 as well as a perdeuterated heparin analog, and measured by NS the dynamical properties of hydration water on the surface of tau monomers and of tau fibers whose formation was triggered by the heparin analog. Both elastic and quasi-elastic NS indicate an increased mobility of hydration water on tau fibers compared with tau monomers. MD simulations provide circumstantial evidence suggesting that it is the increase in water dynamics around the disordered fuzzy coat and not around the fiber core that is at the origin of the experimentally observed increase in tau hydration water dynamics after fibrillation. We conjecture that the observed gain in water dynamics reflects an increase in water entropy that is favorable to the fiber formation.