Mechanisms of amphipathic and cargo delivery peptides

描述（由申请人提供）：本研究的长期目标是开发合理的方法，用于设计和改进膜穿透性、两亲性肽，这些肽是抗生素或细胞溶解性的，或者可以携带其他毒性分子作为货物进入细胞。实现这一目标的关键知识是确定这些肽的膜渗透机制。在获得这些信息的一个重要步骤是在本申请中的实验动力学研究的建议，结合全球的理论分析，我们已经开发，膜渗透由一组已知的细胞毒性或抗菌两亲性肽。已知这类肽表现出相当大的靶特异性，这似乎来源于肽与靶细胞膜的脂质双层的相互作用，而没有蛋白质受体的干预。这种广泛的生物防御系统的机制目前还不清楚。选择长度（14至37个氨基酸）、电荷（0至+6）和特异性（抗生素、抗真菌或溶血）不同的四种肽来测试一组肽机制的当前模型。此外，还将检查嵌合构建体转运蛋白TP 10以确定其作为药物递送载体的机制和有用性。已显示转运蛋白能够将甚至大的蛋白质如绿色荧光蛋白转运到细胞中。将确定货物的附着是否以及如何改变肽作用的机制。为了用作将货物运输到细胞中的载体，肽必须易位到囊泡的内部而不永久插入双层中。本研究将表明哪种类型的肽检查是最好的模板，以建立这样的肽媒介物。载物肽可用于将药物转运到真核细胞中或将抗生素转运到细菌细胞中。克服细胞屏障，包括细胞内区室，是使用抗生素的主要困难。

DESCRIPTION (provided by applicant): The long-term objective of this research is the development of rational methods for the design and improvement of membrane-penetrating, amphipathic peptides that are antibiotic or cytolytic, or can carry other toxic molecules as cargo into cells. Critical knowledge in reaching this objective is the determination of the mechanism(s) of membrane penetration by these peptides. An important step in obtaining this information is the proposal in this application of an experimental kinetic study, combined with a global theoretical analysis that we have developed, of membrane penetration by a set of known cytotoxic or antibacterial amphipathic peptides. Peptides in this class are known to exhibit considerable target specificity which appears to derive from the interaction of the peptides with the lipid bilayer of the target cell membrane without the intervention of protein receptors. The mechanism of this widespread biological defense system is presently not understood. Four peptides that differ in length (from 14 to 37 amino acids), charge (0 to +6), and specificity (antibiotic, antifungal, or hemolytic), were selected to test a set of current models of peptide mechanisms. In addition, the chimeric construct transportan TP10 will also be examined in order to determine its mechanism and usefulness as a drug delivery vehicle. Transportans have been shown to be able to transport even large proteins, such as green fluorescent protein, into cells. It will be determined whether and how attachment of cargo modifies the mechanism of the peptide action. In order to be useful as vehicles to transport cargo into cells, peptides must translocate into the interior of vesicles without getting permanently inserted into the bilayer. This study will indicate which of the types of peptides examined is the best template to build such a peptide vehicle. Cargo-carrying peptides can be used to transport drugs into eukaryotic cells or antibiotics into bacterial cells. Surmounting cellular barriers, including intracellular compartments, is a major difficulty in the use of antibiotics.