Computational Assembly of Beta Barrel Membrane Protein

描述（由申请人提供）：桶状物存在于革兰氏阴性菌、抗酸革兰氏阳性菌、真核线粒体、叶绿体（如大肠杆菌、脑膜炎奈瑟菌、淋病奈瑟菌和分枝杆菌如结核分枝杆菌）的外膜中。革兰氏阳性细菌的许多成孔外毒素也是桶状膜蛋白(例如？-金黄色葡萄球菌的溶血素和炭疽杆菌的保护性抗原)。？桶状膜蛋白在许多基本生物过程中起着重要作用。它们控制离子和有机分子在细菌和线粒体外膜上的交换和运输。除了古生菌外，它们对所有生命领域的蛋白质转运都是必不可少的。它们调节新陈代谢和细胞凋亡。它们在免疫监测、提供抗生素耐药性方面也很重要，并且是细菌毒力的关键决定因素。因此，？桶状膜蛋白是开发感染性疾病药物和疫苗的重要靶点。它们也是开发用于高通量DNA测序的生物纳米孔以及靶向癌症药物递送的纳米设备的重要工程努力的焦点。虽然通过实验和计算研究已经了解了很多，但目前对？-桶膜蛋白是不完整的，只有少数结构是已知的，并且缺乏对？-桶膜蛋白。该研究的长期目标是获得对？的结构、相互作用和功能的基本理解和机制洞察。-桶膜蛋白，并开发使能技术设计？具有增强生物物理特性的桶状膜蛋白。具体目标是：1)开发控制组装的物理原理的计算模型？-桶膜蛋白。将开发粗粒度模型来解释结构稳定性和蛋白质-蛋白质相互作用（PPIs）的关键决定因素。-桶膜蛋白。这将使通过计算对蛋白质稳定性进行定量评估成为可能。2)预测？的结构、寡聚化状态和蛋白质-蛋白质界面-桶膜蛋白。重点将放在预测新建筑结构的挑战性任务上

DESCRIPTION (provided by applicant): Barrels are found in the outer membrane of gram-negative bacteria, acid-fast gram-positive bacteria, eukaryotic mitochondria, chloroplasts (e.g., in E. coli, N. meningitidis, N. gonorrheae, and mycobacteria such as M. tuberculosis). Many pore-forming exotoxins from gram-positive bacteria are also -barrel membrane proteins (e.g., ?-hemolysin of S. aureus and protective antigen of B. anthracis). ?-barrel membrane proteins are important for many fundamental biological processes. They control the ex- change and transport of ions and organic molecules across the bacterial and mitochondrial outer membranes. They are essential for protein translocation in all domains of life, except archaea. They regulate metabolism and apoptosis. They are also important for immune surveillance, in providing resistance to antibiotics, and are key determinants of bacterial virulence. As a result, ?-barrel membrane proteins are important therapeutic targets for developing drugs and vaccines against infectious diseases. They are also the focus of significant engineering efforts in developing biological nanopores for high-throughput DNA sequencing, as well as nano-devices for targeted cancer drug delivery. Although much has been learned through experimental and computational studies, current knowledge of ?-barrel membrane proteins is incomplete Only a small number of structures are known, and there is a lack of understanding of the general organizing principles of ?-barrel membrane proteins. The long term goal of the proposed research is to gain fundamental understanding and mechanistic insight into the structures, interactions, and functions of ?-barrel membrane proteins, and to develop enabling technology for design of ?-barrel membrane proteins with enhanced biophysical properties. The specific aims are to: 1) Develop computational models of physical principles governing the assembly of ?-barrel membrane proteins. Coarse-grained models will be developed to account for key determinants of structural stability and protein-protein interactions (PPIs) of ?-barrel membrane proteins. This will enable quantitative assessment of protein stability through computation. 2) Predict structures, oligomerization state, and protein-protein interfaces of ?-barrel membrane proteins. The focus will be on the challenging tasks of predicting structures of novel architecture or structures with no known templates. In addition, methods will be developed to predict protein oligomerization states and to identify protein-protein interaction sites. Structures with known templates will also be predicted through detection of remote homologs using newly developed technique of evolutionary analysis. 3) Develop engineering principles for designing ?-barrel porins with desirable stability, oligomerization state, and pore geometry. Design strategies for ?-barrel membrane porins with altered oligomerization states and altered stability will be developed. Proteins with enhanced as well as weakened stability, for both monomeric and oligomeric proteins will be designed. In addition, porins with complex pore geometry using naturally occurring building blocks will also be designed. 4) Experimental validation of computational prediction and design. Computational predictions will be verified by experimental studies. Extensive mutant studies will be carried out to test whether designed ?-barrel membrane proteins have the intended changes in stability, in oligomerization state, as well as in geometry.