Study Protein-Nanomaterial Interactions and Their Impacts on Protein Activities

描述（由申请人提供）：由于纳米材料与生物物质的尺寸匹配，它们的大比表面积，以及它们的无机核心材料，纳米材料与生物系统的相互作用不同于小分子或生物聚合物，我们以前知道的物体。随着人类越来越多地以环境污染物或纳米药物的形式接触工程纳米材料，人们迫切需要更好地了解工程纳米材料在生物系统中的行为。一旦ENM进入人体，它们就会遇到大量的蛋白质，并在其表面形成所谓的蛋白质“冕”。ENM的生物学效应可以通过这种蛋白质“冠状”来实现。例如，蛋白质冠可以携带生物系统识别的特殊信号，从而控制ENM的行为，如从循环中清除，渗透到膜屏障，运输到居住地，触发防御系统等。另一方面，吸附在ENM表面的蛋白质可能会钝化其高表面活性并阻止自由基的产生，自由基是纳米毒性的主要来源，但代价是可能改变结合蛋白的构象和活性。通过假设生物系统中蛋白质-纳米材料相互作用与ENM行为之间的直接联系，我们建议分析蛋白质电晕的形成，整理其对ENM物理性质的依赖，并开始探索与ENM结合对蛋白质的潜在影响。我们已经开发了用于相互作用研究的有效工具，包括用于分离高亲和力蛋白质的流场流分馏和用于结合亲和力测量的毛细管电泳。利用这些工具，我们将在人血浆或细胞裂解液中筛选与所选ENM具有高亲和力的蛋白质（Specific Aim 1）。这项研究将揭示ENM进入人体或入侵细胞后冠状细胞中存在的蛋白质。这些蛋白应该在调控生物宿主的ENM行为和宿主对ENM进入的反应中发挥重要作用。同时，由于这些蛋白的密切接触，它们将成为ENM最可能影响的靶点。其次，我们将详细研究一组具有代表性的蛋白质与所选纳米材料之间的相互作用（Specific Aim 2），旨在更好地理解相互作用对ENM性质的一般依赖，如核心材料、尺寸、形状和表面功能化。最后，我们将从酶活性改变的角度评估蛋白质-纳米材料相互作用的结果（Specific Aim 3）。我们的研究将有助于更好地理解纳米材料的生物学效应以及相互作用对ENM性质的一般依赖关系，这可以指导纳米药物的设计，并确保纳米材料的安全实施。

DESCRIPTION (provided by applicant): Due to the matching sizes of nanomaterials with biological substances, their large specific surface areas, and their inorganic core materials, nanomaterials interact with biological systems differently than small molecules or biopolymers, objects we have previous knowledge of. With the increasing exposure of human beings to engineered nanomaterials (ENM) in the form of environmental pollutant or nanomedicine, it is greatly demanded that we obtain better understanding of the behavior of ENM inside the biological systems. Once the ENM enters the human body, they encounter enormous amounts of proteins and form the so-called protein "corona" on their surface. The biological effects of ENM could then be carried out by this protein "corona". For example, the protein corona could bear special signals recognizable by biological systems and thus govern the behaviors of ENM, such as clearance from circulation, penetration to membranous barriers, transportation to residential sites, trigger of defense system, etc. On the other hand, proteins adsorbed onto ENM surface may passivate their high surface activity and prevent generation of free radicals, the main source of nanotoxicity, but at the expense of possible conformational and activity alteration to the bound proteins. By hypothesizing the direct link between protein-nanomaterial interaction and the behavior of ENM in biological systems, we propose to analyze the formation of protein corona, sort out its dependence on the physical properties of ENM, and initiate the exploration of potential effects of binding to ENM on proteins. We have developed effective tools for the interaction study, including flow-field flow fractionation for isolation of proteins of high affinity and capillary electrophoresis for binding affinity measurement. With these tools, we will screen for proteins in human plasma or cell lysate having high affinity to the selected ENM (Specific Aim 1). This study will reveal proteins present in the corona after ENM entering our body or invading cells. These proteins should play important roles in governing the ENM behaviors in their biological hosts and the host response to ENM entry. In the meanwhile, these proteins will be the most possible targets being affected by ENM because of the close contact. Secondly, we will study the interactions between a representative group of proteins and the selected nanomaterials in details (Specific Aim 2), aiming to obtain better understanding on the general dependence of interaction to the ENM properties, such as core materials, size, shape, and surface functionalization. Last, we will assess outcomes of protein-nanomaterial interaction from the aspect of alteration of enzyme activities (Specific Aim 3). Our study will lead to better understanding of the biological effects of nanomaterials and the general dependence of interaction on the ENM properties, which can guide the design of nanomedicines as well as ensure safe implementation of nanomaterials. PUBLIC HEALTH RELEVANCE: We propose experiments to test our hypothesis that the biological behavior of nanomaterials may be carried out via strong interactions with plasma and cellular proteins. Such interactions could alter the protein conformation and thus protein function. The outcome of the propose research may shed light on how the nanomaterials behave inside the biological host, knowledge required by nanomedicine design and nanomaterials exposure control.