Abstract：
The in vivo potency of polyphosphazene immunoadjuvants is inherently linked to the ability of these ionic macromolecules to assemble with antigenic proteins in aqueous solutions and form physiologically stable supramolecular complexes. Therefore, in-depth knowledge of interactions in this biologically relevant system is a prerequisite for a better understanding of mechanism of immunoadjuvant activity. Present study explores a self-assembly of polyphosphazene immunoadjuvant-PCPP and a model antigen-lysozyme in a physiologically relevant environment-saline solution and neutral pH. Three analytical techniques were employed to characterize reaction thermodynamics, water-solute structural organization, and supramolecular dimensions: isothermal titration calorimetry (ITC), water proton nuclear magnetic resonance (wNMR), and dynamic light scattering (DLS). The formation of lysozyme-PCPP complexes at near physiological conditions was detected by all methods and the avidity was modulated by a physical state and dimensions of the assemblies. Thermodynamic analysis revealed the dissociation constant in micromolar range and the dominance of enthalpy factor in interactions, which is in line with previously suggested model of protein charge anisotropy and small persistence length of the polymer favoring the formation of high affinity complexes. The paper reports advantageous use of wNMR method for studying protein-polymer interactions, especially for low protein-load complexes.
摘要：
聚磷腈免疫佐剂的体内效力本质上与这些离子大分子在水溶液中与抗原蛋白组装并形成生理稳定的超分子复合物的能力有关。因此,深入了解该生物学相关系统中的相互作用是更好地了解免疫佐剂活性机制的前提.本研究探索了在生理相关环境-盐溶液和中性pH中，聚磷腈免疫佐剂-PCPP和模型抗原-溶菌酶的自组装。采用三种分析技术来表征反应热力学，水溶质结构组织，和超分子尺寸：等温滴定量热法（ITC），水质子核磁共振（wNMR），和动态光散射(DLS)。通过所有方法检测到在接近生理条件下溶菌酶-PCPP复合物的形成，并且亲和力由组件的物理状态和尺寸调节。热力学分析揭示了微摩尔范围内的解离常数和焓因子在相互作用中的优势，这与先前提出的蛋白质电荷各向异性模型和聚合物的小持久性长度一致，有利于形成高亲和力复合物。本文报道了wNMR方法用于研究蛋白质-聚合物相互作用的有利用途，特别是对于低蛋白负载的复合物。
