Structure, Assembly, and Function of Outer Membrane Proteins

描述（由申请人提供）：膜蛋白约占所有原核和真核蛋白质组的30%。它们负责离子传导、化学运输、能量转换、信号转导、激素和光接收、细胞粘附以及许多其他功能。结构生物学长期坚持的原则是，要充分了解生物分子的功能，就必须知道它们的结构。膜蛋白在这方面也不例外。然而，主要由于技术原因，旨在阐明其结构的科学远远落后于可溶性蛋白质的类似科学。本申请提出的研究目标有两个方面：(1)推进溶液核磁共振光谱作为膜蛋白结构测定的方法；(2)通过溶液核磁共振研究四种原核外膜蛋白的结构，进一步了解它们的功能。具体来说：来自大肠杆菌的OmpA将主要用作核磁共振方法开发的膜蛋白模型，用于研究脂质-蛋白相互作用，以及检查脂质双层中膜蛋白的稳定性；来自大肠杆菌的OmpG正成为生物传感器开发的首选生物纳米孔，核磁共振将用于了解其从打开状态到关闭状态的机制；OprG和OprH是铜绿假单胞菌（Pseudomonas aeruginosa）的两个外膜蛋白，我们将通过确定它们的结构和研究它们的功能来研究它们在铜绿假单胞菌严重耐药中的作用。与公共卫生相关：本项目将推动核磁共振波谱法测定膜结合蛋白的结构和功能的前沿。将被检查的特定膜蛋白在生物传感器开发中具有生物技术应用，或者是铜绿假单胞菌细胞膜的组成部分，即医院引起的感染和囊性纤维化并发症中的临床重要病原体。这项研究可能最终有助于更好地治疗这些感染。

DESCRIPTION (provided by applicant): Membrane proteins constitute about 30 % of all prokaryotic and eukaryotic proteomes. They are responsible for ion conduction, chemical transport, energy conversion, signal transduction, hormone- and photo-reception, cell adhesion, and many other functions. The long-held tenet of structural biology is that to fully understand the function of biomolecules their structure must be known. Membrane proteins are no exception in this regard. However, the science aimed at elucidating their structures has lagged far behind similar science on soluble proteins, mostly for technical reasons. The goal of the research proposed in this application is two-fold: (1) to advance solution NMR spectroscopy as a method for structure determination of membrane proteins and (2) to further the understanding of the functions of four prokaryotic outer membrane proteins by studying their structures by solution NMR. Specifically: OmpA from E. coli primarily will be used as a model membrane protein for NMR methods development, for studying lipid-protein interactions, and for examining the stability of membrane proteins in lipid bilayers; OmpG from E. coli is becoming the preferred biological nanopore for biosensor development and NMR will be used to understand the mechanism, by which it gates from the open to the closed state; and OprG and OprH are two outer membrane proteins from Pseudomonas aeruginosa, whose contributions to the severe antibiotic resistance of this microorganism will be investigated by determining their structures and studying their functions. PUBLIC HEALTH RELEVANCE: This project will push the frontier of determining the structures and understanding the functions of membrane-bound proteins by nuclear magnetic resonance spectroscopy. The specific membrane proteins that will be examined have biotechnological applications in biosensor development or are components of the cell envelope of Pseudomonas aeruginosa, i.e. a clinically important pathogen in hospital-induced infections and complications of cystic fibrosis. The research may ultimately contribute to better treatments of these infections.