Nanoscale organisation of water and ions at bio-interfaces: consequences on anti-infective peptide adsorption

最近的研究表明，水和简单的金属离子可以在溶液中的矿物表面自发地形成相关的、有序的网络。基于原子力显微镜（AFM）的初步结果表明，类似的效应发生在脂质双分子层表面。由于生物界面的软性质，有序界面水/离子结构的存在将对调节生物膜的力学性能、形状和局部流动性产生深远的影响。它还将在介导膜结合分子（如生物标志物、药物和肽）的相互作用以及生物能量学中的电荷转移方面产生影响。本提案利用AFM领域的最新进展来探索界面液体（水和离子）的局部结构与模型膜在经历相变时的行为之间的相互作用。该研究将通过原子力显微镜在溶液中进行，具有分子或单水合离子精度。研究结果将提供界面液体结构的亚纳米图，跟踪它们在膜从凝胶到流体转变过程中的局部演变，确定它们对膜性质（弹性、流动性）的影响，并量化它们在介导抗喷射肽吸附方面的意义。在项目的第一部分，将通过AFM对双层表面的特定区域进行成像和探测（机械性能映射），同时使用精确的环境控制器逐步提高溶液的温度。在整个过程中，包括随后的膜冷却过程，将监测融化膜和界面离子结构对彼此的相互影响。提出的研究考察了膜脂组成和溶液中存在的离子类型等参数对界面结构形成和演化的影响。还将研究涉及不同脂质的二元混合物的界面，以阐明膜物理和化学奇点（如界面过程中的畴边）所起的作用。随后，界面效应对膜结合肽的影响将在生物学相关条件下进行研究。实验将在已知插入膜中的抗菌/抗感染肽（Temporins）存在的情况下进行。目标将是确定在项目前几个阶段观察到的界面效应是否会影响抗菌肽与膜之间的纳米级相互作用，特别是单个肽的结合和随后的插入。从凝胶相膜（无插入）到流体膜，这一过程将被仔细监测，特别注意插入肽的空间位置以及与先前界面奇点的潜在时间相关性。这个项目的发现将是非常新颖的，很可能提供第一个直接观察生物界面中界面介导的过程。

Recent findings have shown that water and simple metal ions can spontaneously create correlated, ordered networks at the surface of minerals in solution. Preliminary results based on atomic force microscopy (AFM) suggest that similar effects occur at the surface of lipid bilayers. Due to the soft nature of biointerfaces, the existence of ordered interfacial water/ion structures would have deep implications for regulating biomembranes' mechanical properties, shape and local fluidity. It would also have implications in mediating the interactions of membrane-binding molecules such as biomarker, drugs and peptides, and for charge transfer in bioenergetics.This proposal exploits recent advances in the field of AFM to explore the interplay between the local structure of the interfacial liquid (water and ions) and the behaviour of model membranes when undergoing a phase transition. The study will be conducted by AFM in solution and with molecular or single hydrated ion precision. The results will provide sub-nanometre maps of interfacial liquid structures, track their evolution locally as the membrane undergoes a gel to fluid transition, establish their influence on the membrane properties (elasticity, fluidity), and quantify their implications in mediating the adsorption of anti-invective peptides. In the first part of the project, specific regions of the bilayer surface will be imaged and probed (mechanical properties mapping) by AFM while the temperature of the solution is progressively increased using a precise environment controller. The mutual effect that the melting membrane and the interfacial ionic structures have on each other will be monitored throughout the process, including upon subsequent cooling of the membrane. The proposed research examines the influence of parameters such as membrane lipid composition and the type of ions present in the solution on the formation and evolution of interfacial structures. Interfaces involving binary mixtures of dissimilar lipids will also be studied in order elucidate the role played by membrane physical and chemical singularities such as domain edges on interfacial processes.Subsequently, the influence of interfacial effects on membrane-binding peptides will be studied in biologically relevant conditions. Experiments will be conducted in the presence of antimicrobial/anti-infective peptides (Temporins) known to insert into the membrane. The goal will be to determine whether the interfacial effects observed in the previous phases of the project can influence the nanoscale interactions between antimicrobial peptides and membranes, in particular the binding and subsequent insertion of single peptides. The process will be carefully monitored, starting from gel-phase membranes (no insertion) to fluid membranes, paying particular attention to the spatial location of inserting peptides and a potential temporal correlation with a prior interfacial singularity. The findings of this project will be highly novel and are likely to provide the first direct observation of interface-mediated processes at biointerfaces.