We investigate the solution and fibril conformations and structural transitions of the polyglutamine (polyQ) peptide, DQK (Q10), by synergistically using UV resonance Raman (UVRR) spectroscopy and molecular dynamics (MD) simulations. We show that Q10 adopts two distinct, monomeric solution conformational states: a collapsed β-strand and a PPII-like structure that do not readily interconvert. This clearly indicates a high activation barrier in solution that prevents equilibration between these structures. Using metadynamics, we explore the conformational energy landscape of Q10 to investigate the physical origins of this high activation barrier. We develop new insights into the conformations and hydrogen bonding environments of the glutamine side chains in the PPII and β-strand-like conformations in solution. We also use the secondary structure-inducing cosolvent, acetonitrile, to investigate the conformations present in low dielectric constant solutions with decreased solvent-peptide hydrogen bonding. As the mole fraction of acetonitrile increases, Q10 converts from PPII-like structures into α-helix-like structures and β-sheet aggregates. Electron microscopy indicates that the aggregates prepared from these acetonitrile-rich solutions show morphologies similar to our previously observed polyQ fibrils. These aggregates redissolve upon the addition of water! These are the first examples of reversible fibril formation. Our monomeric Q10 peptides clearly sample broad regions of their available conformational energy landscape. The work here develops molecular-level insight into monomeric Q10 conformations and investigates the activation barriers between different monomer states and their evolution into fibrils.

Understanding the structure of polyglutamine (polyQ) amyloid-like fibril aggregates is crucial to gaining insights into the etiology of at least ten neurodegenerative disorders, including Huntington's disease. Here, we determine the structure of D2Q10K2 (Q10) fibrils using ultraviolet resonance Raman (UVRR) spectroscopy and molecular dynamics (MD). Using UVRR, we determine the fibril peptide backbone Ψ and glutamine (Gln) side chain χ3 dihedral angles. We find that most of the fibril peptide bonds adopt antiparallel β-sheet conformations; however, a small population of peptide bonds exist in parallel β-sheet structures. Using MD, we simulate three different potential fibril structural models that consist of either β-strands or β-hairpins. Comparing the experimentally measured Ψ and χ3 angle distributions to those obtained from the MD simulated models, we conclude that the basic structural motif of Q10 fibrils is an extended β-strand structure. Importantly, we determine from our MD simulations that Q10 fibril antiparallel β-sheets are thermodynamically more stable than parallel β-sheets. This accounts for why polyQ fibrils preferentially adopt antiparallel β-sheet conformations instead of in-register parallel β-sheets like most amyloidogenic peptides. In addition, we directly determine, for the first time, the structures of Gln side chains. Our structural data give new insights into the role that the Gln side chains play in the stabilization of polyQ fibrils. Finally, our work demonstrates the synergistic power and utility of combining UVRR measurements and MD modeling to determine the structure of amyloid-like fibrils.

We determined an empirical correlation that relates the amide I vibrational band frequencies of the glutamine (Q) side chain to the strength of hydrogen bonding, van der Waals, and Lewis acid-base interactions of its primary amide carbonyl. We used this correlation to determine the Q side chain carbonyl interaction enthalpy (Δ H) in monomeric and amyloid-like fibril conformations of DQK (Q10). We independently verified these Δ H values through molecular dynamics simulations that showed excellent agreement with experiments. We found that side chain-side chain and side chain-peptide backbone interactions in fibrils and monomers are more enthalpically favorable than are Q side chain-water interactions. Q10 fibrils also showed a more favorable Δ H for side chain-side chain interactions compared to backbone-backbone interactions. This work experimentally demonstrates that interamide side chain interactions are important in the formation and stabilization of polyQ fibrils.