CAREER: Using chemistry to probe anthrax toxin protein translocation

有了这个奖项，布拉德利L。来自马萨诸塞州理工学院的Pentelute，由生命过程化学计划资助，将研究蛋白质跨细胞膜易位的机制。自然界已经进化出许多大的蛋白质，其功能是进入哺乳动物细胞。其中一些蛋白质是由细菌产生的，通常对宿主有毒。这种蛋白质的一个例子是炭疽毒素。这项研究的目的是利用化学方法来揭示炭疽毒素蛋白进入细胞的分子基础。一组易位测定和一个蛋白质修饰工具包，允许随意修饰，将用于研究无毒形式的炭疽毒素如何进入细胞。通过采取化学方法，将制备和研究在这些大型炭疽蛋白复合物内的定义位置包含化学修饰的变体，以便对这些纳米机器的操作提供基本的理解。研究生和本科生水平的研究培训将使学生接触这些研究中使用的最新化学方法，并为他们提供对复杂生物过程的洞察力。将进一步努力举办一次外联化学品展览。PI将与麻省理工学院的本科生一起进行化学演示，这些演示对8-12岁的学生来说似乎很神奇。教学理念是，魔术将抓住年轻人的注意力，激发他们对科学的兴趣。本研究计划将系统地揭示货物在跨膜转运中的作用，并设计带电荷的肽，使异源蛋白通过保护性抗原孔。研究程序将采用蛋白质修饰工具包，允许随意修饰和研究炭疽毒素纳米机器。本文的研究将为蛋白质跨膜转运提供一个机制上的理解，并为使用半合成策略来探测转运过程奠定基础。此外，从这些研究中收集到的工具和见解将允许开发抑制剂和方法，将新的化学形式传递到细胞中，这是化学中的一个长期问题。

With this award, Bradley L. Pentelute from the Massachusetts Institute of Technology, funded by the Chemistry of Life Processes Program, will investigate the mechanism of protein translocation across a cell membrane. Nature has evolved a number of large proteins whose function is to enter mammalian cells. Some of these proteins are produced by bacteria and are often toxic to the host. An example of such a protein is anthrax toxin. The aim of the research is to use chemistry to uncover the molecular basis of anthrax toxin protein entry into cells. A battery of translocation assays and a protein modification toolkit, that allows at-will modification, will be used to investigate how the non-toxic forms of anthrax toxin enter cells. By virtue of taking a chemical approach, variants that contain chemical modifications at defined positions within these large anthrax protein complexes will be prepared and investigated in order to provide a fundamental understanding of the operation of these nanomachines. Research training at the graduate and undergraduate levels will expose students to the latest chemical methods that are used in these studies and provide them with insight into complicated biological processes. Additional effort will be put into an outreach chemical show. The PI together with MIT undergraduate students will perform chemical demonstrations that appear to be magical to an audience of 8-12 year old students. The teaching philosophy is that magic will capture the youths' attention and excite them about science. The long-term aim of outreach program is to springboard young patrons into careers of fundamental science.This research project will systematically uncover the role of cargo on translocation across a membrane and design charged peptides for heterologous protein passage through a protective antigen pore. The research procedures will employ a protein modification toolkit that allows at-will modification and investigation of the anthrax toxin nanomachine. The studies proposed here will provide a mechanistic understanding of protein translocation across a membrane and lay the foundation for using semi-synthetic strategies to probe the translocation process. In addition, the tools and insights gleaned from these studies will allow the development of inhibitors and methods to deliver novel chemical forms into cells, which is a longstanding problem in chemistry.