NIH Pioneer Award

描述 摘要： 结晶学在确定数千种不同蛋白质的原子分辨结构方面的成功使生物学发生了革命性的变化。膜蛋白是医学上一个重要的大类，但它没有跟上使这场革命成为可能的方法学进步的步伐。其原因是，与可溶性蛋白质不同，膜蛋白在其生物发生过程中必须进入脂质双层。这种额外的复杂程度阻碍了膜蛋白的获得，其水平与可溶性蛋白相似。结构生物学要取得成功，膜蛋白必须过度表达。问题是，以目前的技术，这对大多数膜蛋白来说是不可能的；因此，确定的结构很少，而且每个结构都非常昂贵。这一建议旨在通过证明以下假设来解决这一问题：膜蛋白过度表达的限制因素是它们需要被靶向并插入膜中。膜蛋白是由N-末端信号识别的，必须准确阅读这些信号才能适当地传递蛋白质，以便通过转位机制将蛋白质插入膜中。我们的目标将是开发一种通用信号，使我们能够克服这一内在的速率限制步骤。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点。通过这种全面的方法，我们将获得生物学上的洞察力；此外，我们将显著增加可用于结晶的膜蛋白靶标的数量。其效果将是提高结晶试验的成功率，显着降低解决膜蛋白结构的成本。希望在膜结构测定方面开启一场新的革命，这将使我们能够解决大量重要的医学问题。我的实验室处于领导这场新革命的独特位置。) 公共卫生相关性： 原子分辨结构证明

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