Folding and Design of Beta Sheets in Membranes

描述（由申请人提供）：整合膜蛋白占所有基因组的约25%。那些使用b-barrel基序的基因组占革兰氏阴性菌基因组的4%，并且也存在于分枝杆菌、衣原体、线粒体和叶绿体中。在包括人类在内的所有生命界中也发现了在膜中自组装成b-片层的外源蛋白质和肽，在那里它们参与病原体毒性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-桶来透化膜。也有许多主要是b-折叠结构的成孔肽抗生素。在这个正在进行的研究计划中，我们正在追求更深入地了解合成和生物膜中b-片层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结构和功能的自组装b-片层肽在膜中，并以确定b-桶膜蛋白，预测其结构和工程师的功能。我们正在使用的方法设计和工程孔形成肽和表征抗菌和细胞毒性活性有潜在的重要的生物技术应用领域的抗生素，生物传感器和药物输送。这些广泛的目标将通过以下试验路线来实现。通过筛选能自组装进入脂质双层膜孔的成员，设计新的成孔肽家族。表征脂质囊泡中的孔形成机制和肽的结构，并交叉表征成孔剂的抗微生物活性。通过筛选库来设计和表征成孔肽的强效、广谱抗菌活性。在基于囊泡的系统中交叉表征抗菌肽，以比较两种系统中结构和功能的决定因素。使用组合化学和高通量筛选设计自组装成跨膜蛋白质样b桶孔的肽。最后，我们将使用基因组学和蛋白质组学来描绘革兰氏阴性菌的b桶蛋白质组，并验证b桶预测算法。这些信息将用于创建一个潜在的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