Department of Chemistry and Applied Biosciences, Swiss Federal Institute of Technology Zurich , Vladimir Prelog Weg 1, 8093, Zurich, Switzerland.
ACS Nano. 2017 Nov 28;11(11):11358-11367. doi: 10.1021/acsnano.7b05895. Epub 2017 Oct 25.
Interactions between proteins and surfaces in combination with hydrodynamic flow and mechanical agitation can often trigger the conversion of soluble peptides and proteins into aggregates, including amyloid fibrils. Despite the extensive literature on the empirical effects of surfaces and mechanical forces on the formation of amyloids, the molecular details of the mechanisms underlying this behavior are still elusive. This limitation is, in part, due to the complex reaction network underlying the formation of amyloids, where several microscopic reactions of nucleation and growth can occur both at the interfaces and in bulk. In this work, we design a high-throughput assay based on nanoparticles and we apply a chemical kinetic platform to analyze the mechanisms underlying the effect of surfaces and mechanical forces on the formation of amyloid fibrils from human insulin under physiological conditions. By considering a variety of polymeric nanoparticles with different surface properties we explore a broad range of repulsive and attractive interactions between insulin and surfaces. Our analysis shows that hydrophobic interfaces induce the formation of amyloid fibrils by specifically promoting the primary heterogeneous nucleation rate. In contrast, mechanical forces accelerate the formation of amyloid fibrils by favoring mass transport and further amplify the number of fibrils by promoting fragmentation events. Thus, surfaces and agitation have a combined effect on the kinetics of protein aggregation observed at the macroscopic level but, individually, they each affect distinct microscopic reaction steps: the presence of interfaces generates primary nucleation events of fibril formation, which is then amplified by mechanical forces. These results suggest that the inhibition of surface-induced heterogeneous nucleation should be considered a primary target to suppress aggregation and explain why in many systems the simultaneous presence of surfaces and hydrodynamic flow enhances protein aggregation.
蛋白质与表面的相互作用，加上流体动力学流动和机械搅拌，往往会触发可溶性肽和蛋白质转化为聚集物，包括淀粉样纤维。尽管有大量关于表面和机械力对淀粉样形成的经验影响的文献，但这种行为背后的分子机制细节仍然难以捉摸。这种局限性部分归因于淀粉样形成的复杂反应网络，其中几个成核和生长的微观反应既可以在界面上发生，也可以在本体中发生。在这项工作中，我们设计了一种基于纳米粒子的高通量测定法，并应用化学动力学平台来分析在生理条件下，表面和机械力对人胰岛素形成淀粉样纤维的影响背后的机制。通过考虑具有不同表面特性的各种聚合物纳米粒子，我们探索了胰岛素与表面之间广泛的排斥和吸引相互作用。我们的分析表明，疏水性界面通过特异性促进初级非均相成核速率来诱导淀粉样纤维的形成。相比之下，机械力通过有利于质量传递来加速淀粉样纤维的形成，并通过促进碎片事件进一步放大纤维的数量。因此，表面和搅拌对宏观水平上观察到的蛋白质聚集动力学有综合影响，但它们各自影响不同的微观反应步骤：界面的存在产生了纤维形成的初级成核事件，然后由机械力放大。这些结果表明，抑制表面诱导的非均相成核应被视为抑制聚集的主要目标，并解释了为什么在许多系统中，表面和流体动力学的同时存在会增强蛋白质聚集。
