Structure-based “rational” drug design strategies fail for diseases associated with intrinsically disordered proteins (IDPs). However, structural disorder allows large-amplitude spontaneous intramolecular dynamics in a protein. We demonstrate a method that exploits this dynamics to provide quantitative information about the degree of interaction of an IDP with other molecules. A candidate ligand molecule may not bind strongly, but even momentary interactions can be expected to perturb the fluctuations. We measure the amplitude and frequency of the equilibrium fluctuations of fluorescently labeled small oligomers of hIAPP (an IDP associated with type II diabetes) in a physiological solution, using nanosecond fluorescence cross-correlation spectroscopy. We show that the interterminal distance fluctuates at a characteristic time scale of 134 ± 10 ns, and 6.4 ± 0.2% of the population is in the “closed” (quenched) state at equilibrium. These fluctuations are affected in a dose-dependent manner by a series of small molecules known to reduce the toxicity of various amyloid peptides. The degree of interaction increases in the following order: resveratrol < epicatechin ∼ quercetin < Congo red < epigallocatechin 3-gallate. Such ordering can provide a direction for exploring the chemical space for finding stronger-binding ligands. We test the biological relevance of these measurements by measuring the effect of these molecules on the affinity of hIAPP for lipid vesicles and cell membranes. We find that the ability of a molecule to modulate intramolecular fluctuations correlates well with its ability to lower membrane affinity. We conclude that structural disorder may provide new avenues for rational drug design for IDPs.

中文翻译：

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自发性波动可以指导结构紊乱蛋白质的药物设计策略

基于结构的“合理”药物设计策略无法解决与内在无序蛋白（IDP）相关的疾病。但是，结构紊乱允许蛋白质中出现大幅度自发的分子内动力学。我们展示了一种利用这种动力学来提供有关IDP与其他分子相互作用程度的定量信息的方法。候选配体分子可能不会牢固结合，但是甚至瞬时相互作用也可以预期扰动波动。我们使用纳秒荧光互相关光谱法，在生理溶液中测量了hIAPP（与II型糖尿病相关的IDP）的荧光标记小寡聚物的平衡波动的幅度和频率。我们显示终端间距离在134±10 ns的特征时间尺度上波动，6.4±0.2％的人口处于平衡的“封闭”（猝灭）状态。这些波动受到一系列已知的降低各种淀粉状蛋白肽毒性的小分子的剂量依赖性影响。相互作用程度按以下顺序增加：白藜芦醇<表儿茶素-槲皮素<刚果红<表没食子儿茶素3-没食子酸酯。这样的排序可以为探索化学空间寻找结合力更强的配体提供指导。我们通过测量这些分子对hIAPP对脂质囊泡和细胞膜亲和力的影响来测试这些测量值的生物学相关性。我们发现分子调节分子内波动的能力与其降低膜亲和力的能力密切相关。