Probing the mechanism of protein export by the bacterial Tat transport system

所有的细菌都会产生在细菌细胞外运作的蛋白质。细菌病原体产生的毒素就是一个很好的例子。因为所有的蛋白质都是在细菌内部制造的，所以细胞外的蛋白质必须穿过通常不透水的细胞膜移出细胞。这项任务是由位于细胞膜上的名为蛋白质转运体的机器执行的。TAT系统在许多不同的细菌中发现，包括那些导致人类疾病的细菌，如结核分枝杆菌(导致结核病)和沙门氏菌(导致食物中毒)。科学家们已经对许多不同的致病细菌证明了TAT系统是它们致病的关键，他们越来越有兴趣开发阻止TAT系统工作的药物。在细菌大肠杆菌中，TAT转运蛋白是一个由三种类型的蛋白质组成的大型蛋白质复合体，分别是TATA、TATB和TATC。这些蛋白质被组织成两个不同的亚复合体。有一个由等量的TatB和TatC组成的亚复合体。这个亚复合体在识别将要跨膜运输的蛋白质方面发挥着非常重要的作用。另一个亚复合体由许多拷贝的塔塔蛋白组成。这在膜上形成了运输蛋白质的通道。为了了解TAT机制是如何工作的，并最终设计出阻止它工作的新药，我们需要知道亚复合体中不同的蛋白质(或亚单位)是如何排列的。这个应用程序旨在了解TatBC子联合体是如何组织的。我们在这个项目中想要回答的一些问题是：(I)TatB和TatC蛋白之间的界面位于哪里？(Ii)在蛋白质通过TAT途径运输过程中，界面如何改变以允许TatB和TatC的动态移动？最后，我们知道，在通过TAT系统运输蛋白质的过程中，TATA和TatBC亚复合体聚集在一起运输蛋白质。我们认为，当这种情况发生时，TATA可能会以与TatB相同的方式与TatC结合，我们希望通过进行类似的实验来测试这一点。

All bacteria produce proteins that operate on the outside of the bacterial cell. A good example of this is the toxins produced by bacterial pathogens. Since all proteins are made inside the bacterium, the extracellular proteins must be moved out of the cell across the normally impermeable cell membrane. This task is carried out by machines termed protein transporters that are located in the cell membrane. The Tat system is found in many different bacteria, including those which cause human diseases such as Mycobacterium tuberculosis (which causes tuberculosis) and Salmonella (which causes food poisoning). Scientists have shown for many different pathogenic bacteria that the Tat system is essential for them to cause disease, and they are becoming increasingly interested in developing drugs that prevent the Tat system from working.In the bacterium E. coli, the Tat transporter is a large protein complex made up of three types of proteins, named TatA, TatB and TatC. These proteins are organised into two different sub-complexes. There is a sub-complex made up of equal amounts of TatB and TatC. This sub-complex plays a very important role in recognising the proteins that are going to be transported across the membrane. The other sub-complex is made up of many copies of the TatA protein. This forms the channel in the membrane through which the proteins are transported.In order to understand how the Tat machinery works, and ultimately to design new drugs to stop it working, we need to know how the different proteins (or subunits) in the sub-complexes are arranged. This application is aimed at understanding how the TatBC sub-complex is organised. Some of the questions we want to answer in this project are (i) where are the interfaces between the TatB and TatC proteins located? (ii) How do the interfaces change to allow dynamic movement of TatB and TatC during the transport of proteins through the Tat pathway? Finally we know that during the transport of proteins through the Tat system the TatA and TatBC subcomplexes come together to transport proteins. We believe that when this happens TatA might bind to TatC in the same way that TatB does, and we would like to test this by carrying out similar experiments.