Molecular interaction between ribosome-inactivating proteins and triterpene saponins

核糖体失活蛋白(RIPs)是一种毒素，它不可逆转地使细胞蛋白质机制失活，从而引发细胞死亡。RIPs是由某些植物产生的，这些植物也产生特殊的低分子糖苷，被称为三萜皂苷。这两种成分都被隔离在植物的种子中。我们已经证明，三七皂甙可以显著增加RIPs的毒性。这一效应依赖于三联皂苷通过生物膜拖拽RIPs的能力，从而导致胞质RIP浓度的增加。在进一步的研究中，我们可以证明三七皂苷的拖拽功能可以用于抗体-RIP偶联和非病毒基因的传递，从而展示了作为一般药物传递平台技术的巨大潜力。相互作用分析表明，三七皂苷与RIPs结合，进一步的生物物理研究表明，三七皂素-RIP-纳米颗粒的形成导致RIPs的膜穿透能力增强。据推测，这是通过形成包裹RIPs的三七皂素胶束来促进的。已知三七皂甙可以形成胶束，而胶束又可以穿透生物膜。本项目的目的是研究三七皂苷和RIPs之间的分子相互作用。为此目的，将在电子计算机中执行方法以鉴定RIP上的三萜皂苷的结合位置。通过突变方法，适当的结合位点将逐步改变为非结合位点。通过基于细胞和生物物理的方法，研究这些结合位点对拖拽功能和复合体形成的意义。该项目的结果提供的基本见解可能是开发新的创新策略以增加药物在细胞内传递的起点。

Ribosome-inactivating proteins (RIPs) are toxins that irreversibly inactivate the cellular protein machinery thus triggering cell death. RIPs are produced by certain plants that do also produce particular low molecular glycosides, which are termed triterpensaponins. Both components are sequestered within the plants’ seeds. We have shown that triterpensaponins massively increase the toxicity of RIPs. This effect relies on the ability of the triterpensaponins to drag the RIPs through biological membranes resulting in an increase of the cytosolic RIP concentration. In further studies we could demonstrate that the dragger function of triterpensaponins may be utilized for antibody-RIP-conjugates and non-viral gene delivery thus demonstrating the tremendous potential as platform technology for drug-delivery in general.Interaction analysis have shown that triterpensaponins bind to RIPs and further biophysical investigations indicate the formation of nanoparticulate triterpensaponin-RIP-complexes leading to an enhanced membrane penetration of the RIPs. It is hypothesized that this is facilitated by the formation of triterpensaponin-micelles encapsulating RIPs. Triterpensaponins are known to form micelles and micelles are in turn known to penetrate through biological membranes. The aim of the current project is to investigate the molecular interaction of triterpensaponins and RIPs. For this purpose in silico methods will be performed in order to identify the binding sites of the triterpensaponins on the RIP. By mutational approaches the appropriate binding sites will be than stepwise changed to non-binding sites. By using cell- and biophysical based methods the significance of these binding sites for the dragging function and the complex formation will be investigated.The basic insights provided by the results of this project might be the starting point for the development of novel innovative strategies to increase the intracellular delivery of drugs.