Supramolecular Tandem Membrane Assays

有机分子的膜转运在小分子药物的发现和递送以及抗菌肽和细胞穿透肽（CPP）的作用机制中起着至关重要的作用。然而，它仍然是一个主要的实验挑战，以测量这些化合物的实际运输通过脂质双层膜。我们最近已经表明，脂质体封装的超分子报告对优秀的资格明确检测膜转运的有机分子和肽通过一个简单的和基于荧光的方法。在这个项目中，我们希望进一步建立这种方法的实用性和适用性，因此，我们将在不同大小和膜组成的脂质体中封装超分子报告对，我们将通过吸收和荧光光谱方法研究选定的有机分子和肽的膜转运。这包括光谱表征、时间分辨测量（包括用于快速动力学的停流技术），以及使用巨型单层囊泡（GUV）的荧光显微镜。我们将进一步开发这些“超分子串联膜测定”，将其应用于跟踪小分子的膜运输，未标记分子和CPP。详细地说，我们计划建立膜运输过程的机械调查的方法，测量快速外排动力学，我们将结合联合收割机串联膜检测与酶促反应，为超生物分子传感器系统的发展提供新的途径。

Membrane transport of organic molecules plays a crucial role in the discovery and delivery of small-molecule drugs as well as in the mechanism of action of antimicrobial peptides and cell-penetrating peptides (CPPs). It still remains, however, a major experimental challenge to measure the actual transport of these compounds across the lipid bilayer membrane. We have recently shown that liposome-encapsulated supramolecular reporter pairs excellently qualify to unambiguously detect membrane transport of organic molecules and peptides by a simple and fluorescence-based method. Within this project, we want to further establish the utility and applicability of this method.Therefore, we will encapsulate supramolecular reporter pairs in liposomes of varying size and membrane composition, and we will investigate the membrane transport of selected organic molecules and peptides by absorption and fluorescence spectroscopic methods. This includes spectroscopic characterisations, time-resolved measurements including stopped-flow techniques for rapid kinetics, as well as fluorescence microscopy with giant uni-lamellar vesicles (GUVs).We will further develop these “supramolecular tandem membrane assays” by applying them to follow membrane transport of small, unlabeled molecules and CPPs. In detail, we plan to establish multicolor methods for mechanistic investigation of membrane transport processes, measure fast efflux kinetics, and we will combine tandem membrane assays with enzymatic reactions to provide new avenues for the development of supra-biomolecular sensor systems.