Role of water in protein aggregation and amyloid polymorphism

Abstract

A variety of neurodegenerative diseases are associated with amyloid plaques, which begin as soluble protein oligomers but develop into amyloid fibrils. Our incomplete understanding of this process underscores the need to decipher the principles governing protein aggregation. Mechanisms of in vivo amyloid formation involve a number of coconspirators and complex interactions with membranes. Nevertheless, understanding the biophysical basis of simpler in vitro amyloid formation is considered important for discovering ligands that preferentially bind regions harboring amyloidogenic tendencies. The determination of the fibril structure of many peptides has set the stage for probing the dynamics of oligomer formation and amyloid growth through computer simulations. Most experimental and simulation studies, however, have been interpreted largely from the perspective of proteins: the role of solvent has been relatively overlooked in oligomer formation and assembly to protofilaments and amyloid fibrils. In this Account, we provide a perspective on how interactions with water affect folding landscapes of amyloid beta (Aβ) monomers, oligomer formation in the Aβ16-22 fragment, and protofilament formation in a peptide from yeast prion Sup35. Explicit molecular dynamics simulations illustrate how water controls the self-assembly of higher order structures, providing a structural basis for understanding the kinetics of oligomer and fibril growth. Simulations show that monomers of Aβ peptides sample a number of compact conformations. The formation of aggregation-prone structures (N*) with a salt bridge, strikingly similar to the structure in the fibril, requires overcoming a high desolvation barrier. In general, sequences for which N* structures are not significantly populated are unlikely to aggregate. Oligomers and fibrils generally form in two steps. First, water is expelled from the region between peptides rich in hydrophobic residues (for example, Aβ16-22), resulting in disordered oligomers. Then the peptides align along a preferred axis to form ordered structures with anti-parallel β-strand arrangement. The rate-limiting step in the ordered assembly is the rearrangement of the peptides within a confining volume. The mechanism of protofilament formation in a polar peptide fragment from the yeast prion, in which the two sheets are packed against each other and create a dry interface, illustrates that water dramatically slows self-assembly. As the sheets approach each other, two perfectly ordered one-dimensional water wires form. They are stabilized by hydrogen bonds to the amide groups of the polar side chains, resulting in the formation of long-lived metastable structures. Release of trapped water from the pore creates a helically twisted protofilament with a dry interface. Similarly, the driving force for addition of a solvated monomer to a preformed fibril is water release; the entropy gain and favorable interpeptide hydrogen bond formation compensate for entropy loss in the peptides. We conclude by offering evidence that a two-step model, similar to that postulated for protein crystallization, must also hold for higher order amyloid structure formation starting from N*. Distinct water-laden polymorphic structures result from multiple N* structures. Water plays multifarious roles in all of these protein aggregations. In predominantly hydrophobic sequences, water accelerates fibril formation. In contrast, water-stabilized metastable intermediates dramatically slow fibril growth rates in hydrophilic sequences.

Fig. 1

Schematic of protein aggregation mechanisms leading to polymorphic fibrils. On the left are solvated peptides. Water in the hydration layer is in red and the bulk water in blue. Even isolated monomers sample aggregation prone conformations, N*, which are coated with varying number of water molecules. The peptides with N* conformations aggregate to form disordered protein rich droplets. A major driving force for aggregation is the release of water molecules in the hydration layer into the bulk, which facilitates fibril formation entropically favorable. The structured protein aggregates nucleate from the protein rich droplet to form protofilaments, which further self-assembles to form a variety of mature amyloid fibrils. In some of the polymorphic structures discrete number of water molecules are confined in the fibril.

Fig. 2

Folding landscape of Aβ_10–35 monomers. (A) Low free energy conformations in the which D23 and K28 amino acids, which forms a salt bridge in the fibril, are separated by three, two and one water solvation shells respectively (from top to bottom). The backbone oxygen and nitrogen atoms are in red and blue, respectively. The positively and negatively charged, polar, and hydrophobic residues are colored blue, red, purple, and green, respectively. Water molecules around D23 and K28 are in cyan, while water molecules which separate the two residues are shown in yellow. Hydrogen bonds are shown as black dashed lines. (B) Hairpin-like conformation of the Aβ_10–35 monomer which has a topologically similar structure as the peptide structure in the Aβ fibrils. The D23-K28 salt bridge is solvated by the water molecules. The driving force for the formation of hairpin-like conformation is the interaction between the hydrophobic residues in the N and C termini shown in red and green respectively.

Fig. 3

Water release in fibril growth. (A) Variation in the number of water molecules (red) within 3.5Å of the peptide from Sup35, which docks and locks onto the fibril as a function of time. Time-dependent changes (green) in the number of water molecules in the neighborhood of the fibril monomer onto which the solvated peptide docks. (B) Release of water molecules in the zipper region of the fibril occurs in two stages. In the first stage, water is eliminated rapidly as the peptide docks onto the fibril, while in the second stage, the last two water molecules are squeezed out with the concurrent formation of the protofilament with a dry interior (structure on the right).

Fig. 4

Water molecules play a central role in the association kinetics of two sheets formed from peptides rich in amino acids with polar side chains. In the association process starting from a fully solvated pore (structure on the left) trapped water molecules between the protofilaments form ordered water wires (top middle structure). If the sheets misalign confined water molecules are disordered. Release of trapped water molecules results in protofilament formation (structure on the right). In the upper pathway the water molecules in the wire file out in orderly fashion whereas in the bottom pathway water escapes from the crevice on the sides of the protofilament.