Monomeric Polyglutamine Structures That Evolve into Fibrils

We investigate the solution and fibril conformations and structural transitions of the polyglutamine (polyQ) peptide, D2Q10K2 (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.

本研究结合紫外共振拉曼（UV resonance Raman, UVRR）光谱与分子动力学（molecular dynamics, MD）模拟，对聚谷氨酰胺（polyglutamine, polyQ）肽D2Q10K2（简称Q10）的溶液构象、原纤维构象及其结构转变展开系统探究。研究发现，Q10存在两种截然不同的单体溶液构象态：塌陷β折叠链与聚脯氨酸II（PPII）样结构，二者难以发生相互转换。这一结果清晰表明，溶液中存在较高的活化能垒，阻碍了这些构象间的平衡过程。通过元动力学（metadynamics）方法，我们解析了Q10的构象能量景观，以探究该高活化能垒的物理起源。本研究针对溶液中PPII样与β折叠样构象内谷氨酰胺侧链的构象特征与氢键环境提出了全新认知。我们还利用可诱导二级结构形成的助溶剂乙腈，探究了溶剂-肽氢键减弱的低介电常数溶液中的肽构象变化。随着乙腈摩尔分数的升高，Q10逐渐从PPII样结构转变为α螺旋样结构与β sheet聚集体。电子显微镜观测结果显示，从富乙腈溶液中制备的聚集体，其形貌与本团队此前观测到的polyQ原纤维高度相似。这些聚集体在加水后可完全重新溶解，这是可逆原纤维形成的首个实证案例。本研究中的单体Q10肽可清晰采样其构象能量景观中的广泛区域。本工作从分子层面深入解析了单体Q10的构象特征，并探究了不同单体构象间的活化能垒及其向原纤维演化的过程。