Abstract：
Reflectin is an intrinsically disordered protein (IDP) known for its ability to modulate the biophotonic camouflage of cephalopods based on its assembly-induced osmotic properties. Its reversible self-assembly into discrete, size-controlled clusters and condensed droplets are known to depend sensitively on the net protein charge, making reflectin stimuli-responsive to pH, phosphorylation, and electric fields. Despite considerable efforts to characterize this behavior, the detailed physical mechanisms of reflectin\'s assembly are yet fully understood. Here, we pursue a coarse-grained molecular understanding of reflectin assembly using a combination of experiments and simulations. We hypothesize that reflectin assembly and phase behavior can be explained from a remarkably simple colloidal model whereby individual protein monomers effectively interact via a short-range attractive and long-range repulsive (SA-LR) pair potential. We parameterize a coarse-grained SA-LR interaction potential for reflectin A1 from small angle X-ray scattering measurements, and then extend it to a range of pH using Gouy-Chapman theory to model monomer-monomer electrostatic interactions. The pH-dependent SA-LR interaction is then used in molecular dynamics simulations of reflectin assembly, which successfully capture a number of qualitative features of reflectin, including pH-dependent formation of discrete-sized nanoclusters and liquid-liquid phase separation at high pH, resulting in a putative phase diagram for reflectin. Importantly, we find that at low pH, size-controlled reflectin clusters are equilibrium assemblies, which dynamically exchange protein monomers to maintain an equilibrium size distribution. These findings provide a mechanistic understanding of the equilibrium assembly of reflectin, and suggest that colloidal-scale models capture key driving forces and interactions to explain thermodynamic aspects of native reflectin behavior. Furthermore, the success of SA-LR interactions presented in this study demonstrates the potential of a colloidal interpretation of interactions and phenomena in a range of IDPs.
摘要：
Reflecin是一种固有的无序蛋白（IDP），以其基于其组装诱导的渗透特性来调节头足类动物的生物光子伪装的能力而闻名。其可逆的自组装成离散的，已知大小控制的簇和凝聚的液滴敏感地依赖于净蛋白质电荷，使反射素刺激响应pH，磷酸化,和电场。尽管做出了相当大的努力来描述这种行为，反射组件的详细物理机制尚未完全理解。这里,我们追求一个粗粒度的分子理解反射组装使用实验和模拟相结合。我们假设反射组装和相行为可以从一个非常简单的胶体模型中解释，其中单个蛋白质单体通过短程吸引和长程排斥（SA-LR）对电势有效地相互作用。我们从小角度X射线散射测量参数化了反射A1的粗粒度SA-LR相互作用电位，然后使用Gouy-Chapman理论将其扩展到pH范围，以模拟单体-单体的静电相互作用。然后将pH依赖性SA-LR相互作用用于反射组装的分子动力学模拟，它成功地捕获了反射的许多定性特征，包括pH依赖性的离散大小的纳米团簇的形成和在高pH下的液-液相分离，产生了一个推定的反射相图。重要的是,我们发现在低pH下，尺寸控制的反射团簇是平衡组件，动态交换蛋白质单体以保持平衡的大小分布。这些发现提供了对反射素平衡组装的机械理解，并建议胶体尺度模型捕获关键驱动力和相互作用来解释天然反射行为的热力学方面。此外，本研究中SA-LR相互作用的成功证明了对一系列IDP中的相互作用和现象进行胶体解释的潜力。
