NIH Pioneer Award

描述 摘要： 晶体学在确定数千种不同蛋白质的原子分辨率结构方面的成功已经彻底改变了生物学。膜蛋白，一个大的和医学上重要的类，没有跟上的进步，使这场革命成为可能的方法。其原因是，与可溶性蛋白质不同，膜蛋白在其生物合成过程中必须进入脂质双层。这种额外的复杂性水平阻止了以与可溶性蛋白相似的水平获得膜蛋白。为了使结构生物学成功，膜蛋白必须过表达。问题是，以目前的技术，这对大多数膜蛋白是不可能的;因此，很少有结构被确定，每一个都非常昂贵。该提案旨在通过证明以下假设来解决这个问题：膜蛋白过表达的限制因素是它们需要被靶向并插入膜中。膜蛋白由N-末端信号识别，其必须被准确地读取以适当地递送蛋白以经由易位机制插入膜中。我们的目标将是开发一种通用的信号，使我们能够克服这种内在的限速步骤。我们将通过系统地测试膜蛋白靶向的各个方面来实现这一目标。通过这种全面的方法，我们将获得生物学见解;此外，我们将大大提高可用于结晶的膜蛋白靶点的数量。其效果将是提高结晶试验的成功率，大大降低解决膜蛋白结构的成本。我们希望在膜结构测定方面开启一场新的革命，使我们能够解决大量重要的医学问题。我的实验室是领导这场新革命的唯一位置。) 公共卫生相关性： 原子分辨率结构证明

DESCRIPTION Abstract: The success of crystallography in determining atomic resolution structures of thousands of diverse proteins has revolutionized biology. Membrane proteins, a large and medically important class, have not kept pace with the advances in methodology that have made this revolution possible. The reason for this is that, unlike soluble proteins, membrane proteins must enter into the lipid bilayer during their biogenesis. This additional level of complexity has prevented membrane proteins from being obtained at levels similar to soluble proteins. For structural biology to be successful, a membrane protein must be overexpressed. The problem is that with current technology this has been impossible for the majority of membrane proteins; therefore, few structures are determined and each is extremely expensive. This proposal aims to solve this problem by proving the following hypothesis: The limiting factor in membrane protein overexpression is their need to be targeted and inserted into the membrane. Membrane proteins are recognized by N-terminal signals that must be accurately read to appropriately deliver the protein for insertion into the membrane via the translocation machinery. Our goal will be to develop a universal signal that will allow us to overcome this intrinsic rate- limiting step. We will accomplish this by systematically testing every aspect of membrane protein targeting. With this comprehensive approach, we will derive biological insight; moreover, we will dramatically improve the number of membrane protein targets available for crystallization. The effect will be to improve the success rate of crystallization trials dramatically reducing the cost of solving a membrane protein structure. The hope is to open up a new revolution in membrane structure determination that will allow us to address a wealth of important medical problems. My laboratory is uniquely positioned to lead this new revolution. ) Public Health Relevance: Atomic resolution structures prov