Interaction of insecticidal toxin proteins with target membranes

在农业中，杀虫蛋白在控制损害农作物和降低产量的害虫物种方面越来越重要。许多结构不同的蛋白质家族能够选择性地裂解目标昆虫细胞，使它们既可以安全地供人类使用/消费，也可以安全地用于非目标物种，包括有益的昆虫。深入了解这些蛋白质的作用机制对于它们的发育和解决因抗虫性而可能出现的问题非常重要。该项目将利用最先进的方法和内部开发的新型显微镜技术相结合，研究杀虫成孔蛋白与靶膜的生物物理和生化相互作用。重组蛋白将被表达和纯化，以便在从化学成分受控的人造脂双层到细胞膜的日益复杂的测试系统中进行分析。将使用最先进的设备(电生理学、荧光显微镜)研究孔的形成，以表征孔的性质及其对膜成分和物理条件(如温度、pH)的依赖。在生物化学和物理之间的交界处，将使用内部开发的尖端无标记光学显微镜技术，以亚毫秒的时间分辨率在纳米级分析单个蛋白质与类脂膜的详细相互作用，而不引入结构-功能人工制品。这将使我们能够回答关键问题，包括i)蛋白质在空间和时间上如何在膜内重塑和扩散，ii)蛋白质如何以及在哪里分配，这取决于不同种类的脂膜的化学组成和曲率，iii)蛋白质的功能是如何受脂环境调节的，以及脂膜的局部组成和曲率是如何受蛋白质影响的(这是一个经常被忽视的相互作用)。值得注意的是，细菌、昆虫和哺乳动物的细胞膜化学成分非常不同。这在调节毒素特异性方面可能很重要，这一方面尚未得到很好的研究，本项目将专门探索这一方面。蛋白质-膜相互作用研究将通过对形成孔的毒素的结构和功能的调查来补充，可能包括结构分析(结晶学、电子显微镜、建模)和诱变研究。研究环境：学生将加入一个充满活力的多学科研究团队，在生命科学/物理界面。主导PI研究杀虫毒素的结构/功能，而量子光电子学和生物光子学小组的Co-PI则利用从固体物理到生物成像和生物传感的一系列主题的技术。学生将参加定期会议，小组成员将讨论他们的研究，并将被鼓励参加物理学院和生物科学学院每周的研讨会。

Insecticidal proteins are of increasing importance in agriculture to control pest species that damage crops and reduce yields. Many structurally-distinct families of proteins are able to lyse target insect cells in a selective manner, making them both safe in terms of human use/consumption and safe for non-target species, including beneficial insects. An in depth understanding of the mechanisms of action of these proteins is important for their development and to tackle problems that may arise due to insect resistance. This project will study the biophysical and biochemical interactions of insecticidal pore-forming proteins with target membranes, using a combination of state-of-the art methods and novel microscopy techniques developed in-house. Recombinant proteins will be expressed and purified for analysis in test systems of increasing complexity from artificial lipid bilayers with controlled chemical composition to cell membranes. The formation of pores will be studied using state of the art equipment (electrophysiology, fluorescence microscopy) to characterise pore properties and their dependence on membrane composition and physical conditions (eg temperature, pH). At the interface between biochemistry and physics, cutting edge label-free optical microscopy techniques developed in-house will be used to analyse the detailed interaction of individual proteins with lipid membranes at the nanoscale with sub-millisecond time resolution, without introducing structural-functional artefacts. This will enable us to answer key questions including i) how proteins remodel and diffuse within membranes in space and time, ii) how and where proteins partition, depending on the heterogeneous lipid membrane chemical composition and curvature, iii) how is the protein function modulated by the lipid environment and how is the lipid membrane local composition and curvature affected by the protein (an interplay often overlooked). Notably, bacteria, insects and mammals have very different cell membrane chemical compositions. This is likely to be important in regulating toxin specificity, an aspect not yet well studied that will be specifically explored in this project.Protein-membrane interaction studies will be complemented by investigations on the structure and function of the pore forming toxins, that may include structural analysis (crystallography, electron microscopy, modelling) and mutagenic studies.Research Environment: The student will join a vibrant multi-disciplinary research team at the life science/physics interface. The lead PI studies the structure/function of insecticidal toxins while the Co-PIs in the Quantum Optoelectronics and Biophotonics group utilise techniques across a range of topics, from solid state physics to bio-imaging and biosensing. The student will join regular meetings where group members will discuss their research and will be encouraged to attend to weekly seminars at the School of Physics and School of Biosciences.