ORIENTED PROTEIN ARRAYS ON POLYMERS NANOSTRUCTURES

提出了一种新的技术，用于设计固定在聚合物纳米球上的二维、定向和功能蛋白质阵列，通过适当功能化的脂质体作为模板。细胞色素c(Cytc)通过电荷残基斑块吸附在阴离子脂双分子膜上，在电子转移过程中负责前驱复合体的形成。结合到大的单层囊泡内壁的Cyt c阵列将通过包裹的预聚体的光引发聚合和蛋白质与聚合物基质的共价交联来固定化。去除脂质双层将释放纳米球，并暴露出功能蛋白质表面积。这些细胞色素c纳米结构的生物活性将通过各种方法来确定，这些方法还将探索它们作为电子转移生物传感器的介体和电子转移蛋白的亲和探针的适用性，以及它们为调节生物电子转移所必需的大分子识别过程提供新的洞察力的潜力。本文提出的固定化技术可以应用于其他由于配体结合或静电相互作用而在具有特定取向的脂质双层上自组装的蛋白质。这是一种简单、通用的方法来获得功能定向的蛋白质阵列。

A novel technique is presented for engineering two dimensional, oriented, and functional protein arrays immobilized on polymer nanospheres, templated through appropriately functionalized liposomes. Cytochrome c (cyt c) adsorbs on anionic lipid bilayers via the patch of charged residues responsible for precursory complex formation during electron transfer. Cyt c arrays bound to the inside wall of large unilamellar vesicles will be immobilized through photo initiated polymerization of an encapsulated prepolymer and covalent crosslinking of the protein to the polymer matrix. Removal of the lipid bilayer will release the nanosphere and expose the functional protein surface area. The biological activity of these cyt c nanostructures will be established through a variety of methods, which will also explore their applicability as mediators in electron transfer biosensors, and as affinity probes for electron transfer proteins, and their potential to provide new insight into the macromolecular recognition processes essential for regulating biological electron transfer. The immobilization techniques presented herein can be applied to other proteins which self- assemble on lipid bilayers with specific orientation due to ligand-binding or electrostatic interactions. It is a simple, general method to obtain functional, oriented protein arrays.