Folding and Design of Beta Sheets in Membranes

描述（由申请人提供）：完整膜蛋白约占所有基因组的25%。使用b桶基序的基因占革兰氏阴性菌基因组的4%，也存在于分枝杆菌、衣原体、线粒体和叶绿体中。在包括人类在内的所有生命体中也发现了在膜中自组装成b片的外源性蛋白质和肽，它们参与病原体毒力和宿主防御以及许多毒素和毒液的作用。例如，炭疽毒素和金黄色葡萄球菌的a-溶血素使用b-桶来渗透膜。也有许多以b-sheet结构为主的成孔肽抗生素。在这个正在进行的研究项目中，我们正在深入了解b-sheet在合成膜和生物膜中自组装的驱动力和结构-功能关系，以设计具有有趣和有用的膜结构和功能的自组装b-sheet肽，并鉴定b-barrel膜蛋白，预测其结构和设计其功能。我们用于设计和工程成孔肽以及表征抗菌和细胞毒性活性的方法在抗生素，生物传感器和药物传递领域具有潜在的重要生物技术应用。这些广泛的目标将通过以下试验来实现。通过筛选在脂质双层膜中自组装成孔的成员的文库，设计新的孔形成肽家族。表征脂质囊泡的孔隙形成机制和肽的结构，并交叉表征孔隙形成物的抗菌活性。设计和表征成孔肽筛选文库有效，广谱抗菌活性。交叉表征抗菌肽在囊泡为基础的系统比较结构和功能的决定因素在两个系统。使用组合化学和高通量筛选设计肽自组装成膜跨越，蛋白质样的b桶孔。最后，我们将使用基因组学和蛋白质组学来描述革兰氏阴性菌的b桶蛋白质组，并验证b桶预测算法。这些信息将用于创建潜在外膜蛋白的公共数据库

DESCRIPTION (provided by applicant): Integral membrane proteins comprise about 25% of all genomes. Those that use the b-barrel motif comprise up to 4% of the genomes of Gram-negative bacteria, and are also present in mycobacteria, chlamydiae, mitochondria and chloroplasts. Exogenous proteins and peptides that self-assemble into b-sheets in membranes are also found in all kingdoms of life, including humans, where they are involved in pathogen virulence and host defense and in the action of many toxins and venoms. For example, the Anthrax toxin and the a-hemolysin of Staphylococcus aureus use b-barrels to permeabilize membranes. There are also many pore-forming peptide antibiotics that are predominantly b-sheet structure. In this ongoing research program we are pursuing a deeper understanding of the driving forces and structure-function relationships for the self-assembly of b-sheets in synthetic and biological membranes in order to design self-assembling b-sheet peptides that have interesting and useful structures and functions in membranes, and in order to identify b-barrel membrane proteins, predict their structure and engineer their function. The methods we are using for designing and engineering pore-forming peptides and for characterizing antimicrobial and cytotoxic activity have potentially important biotechnology applications in the fields of antibiotics, biosensors and drug delivery. These broad goals will be accomplished through the following lines of experimentation. Design new families of pore-forming peptides by screening libraries for members that self-assemble into pores in lipid bilayer membranes. Characterize the mechanism of pore formation in lipid vesicles and the structure of the peptide and cross characterize the antimicrobial activity of pore formers. Design and characterize pore-forming peptides by screening libraries for potent, broad-spectrum antimicrobial activity. Cross characterize antimicrobial peptides in vesicle-based systems to compare the determinants of structure and function in the two systems. Use combinatorial chemistry and high throughput screening to design peptides that self-assemble into membrane-spanning, protein-like b-barrel pores. Finally we will use genomics and proteomics to delineate the b-barrel proteomes of Gram-negative bacteria and validate b-barrel prediction algorithms. This information will be used to create a public database of potential outer membrane proteins