Structure, Orientation, and Competitive Interactions of S. Epidermidis Biofilm Proteins on Surfaces

生物膜形成的第一步是细菌附着到表面。这种连接是由 细胞表面上的成分以及表面本身。了解所涉及的化学相互作用 是预防生物膜相关疾病的重要部分。几种蛋白质与 细菌附着和生物膜起始，但它们在表面上的行为知之甚少。此外，少数 存在能够表征表面结合蛋白行为的实验技术。这个项目 研究了表皮链球菌中两种生物膜相关蛋白Aap和BAE的特性，因为它们 与表面相互作用。最近开发的方法，使用核磁共振光谱将被用来研究 这些蛋白质在各种纳米颗粒表面上的结构和取向。纳米颗粒提供了一个重要的 与宏观表面相比，表面与体积比增加，最近的证据表明， 纳米颗粒曲率基本上不改变蛋白质-表面相互作用的性质。这使得 纳米粒子是研究蛋白质-表面相互作用的一个有吸引力的系统。在这项工作中， 用于模拟由玻璃、塑料和钛（通常用于医疗器械）制成的材料表面， 以及暴露于宿主细胞外基质蛋白的表面。提出了三个具体目标：（1） 确定生物膜相关蛋白质和表面之间相互作用的结构模式，基于NMR 氢氘交换、化学标记和弛豫测量将用于表征 结合到表面上的Aap和CNOE结构域。(2)为了了解生物膜蛋白竞争如何影响 表面结合，将研究Aap和CNOE结构域的混合物，监测同时的表面结合 在真实的时间里。(3)为了确定生物膜蛋白如何与化学钝化表面结合，我们将 探索调节与表面结合的Aap和BAE结构域的蛋白质特性（例如电荷、大小） 用PEG和Tween-20包被。用PEG处理是减少蛋白质结合的常用策略，但 这并不完全妨碍约束力，原因尚不清楚。生物膜是导致 医院相关感染在美国，这个项目将导致更好地了解早期阶段， 细菌附着该项目采用新颖和创新的技术来研究 Aap和BAE的吸附，并且结果也将与其他细菌生物膜直接相关。的 该项目揭示的机制细节将有助于理解生物膜如何形成， 可能最终导致更好的方法来抑制表面上生物膜的形成。

The first step in biofilm formation is bacterial attachment to a surface. This attachment is mediated by components on the cell surface as well as the surface itself. Understanding the chemical interactions involved in attachment is an important part of preventing biofilm-related illness. Several proteins have been implicated in bacterial attachment and biofilm initiation, but their behavior on surfaces is poorly understood. Moreover, few experimental techniques exist that are able to characterize surface-bound protein behavior. This project investigates the properties of two biofilm-related proteins in Streptococcus epidermidis, Aap and AtlE, as they interact with surfaces. Recently developed approaches using NMR spectroscopy will be employed to study the structure and orientation of these proteins on various nanoparticle surfaces. Nanoparticles offer a significant increase in surface to volume ratio compared to macroscopic surfaces, and recent evidence suggests that nanoparticle curvature does not substantially alter the nature of protein-surface interactions. This makes nanoparticles an attractive system for studying protein-surface interactions. In this work, nanoparticles are used to model surfaces of materials made of glass, plastic, and titanium (commonly used in medical devices), as well as surfaces exposed to host extracellular matrix proteins. Three specific aims are proposed: (1) To identify the structural mode of interaction between biofilm-related proteins and surfaces, NMR-based hydrogen deuterium exchange, chemical labeling, and relaxation measurements will be used to characterize Aap and AtlE domains bound to surfaces. (2) To understand how biofilm protein competition influences surface binding, mixtures of Aap and AtlE domains will be studied, monitoring simultaneous surface binding in real time. (3) To determine how biofilm proteins bind to chemically passivated surfaces, we will explore the protein properties (e.g. charge, size) of Aap and AtlE domains that modulate binding to surfaces coated with PEG and Tween-20. Treatment with PEG is a common strategy for reducing protein binding, but this does not prevent binding entirely, and the reason why is not clear. Biofilms represent a major cause of hospital-associated infection in the US, and this project will lead to a better understanding of the early stages of bacterial attachment. This project applies novel and innovative techniques to study the chemical basis of adsorption of Aap and AtlE, and the results will be directly relevant to other bacterial biofilms as well. The mechanistic details revealed by this project will be useful in understanding how biofilms form, and such insights could ultimately lead to better approaches for inhibiting the formation of biofilms on surfaces.