Structure and dynamics of bacterial membrane protein - receptor interactions

细菌膜蛋白-受体相互作用的结构和动力学

基本信息

批准号：
8212381
负责人：
Linda M Columbus
金额：
$ 30.41万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-04-01 至 2014-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8212381
关键词：
Affinity Antibiotics Bacteria Bacterial Outer Membrane Proteins Binding CEACAM1 Caliber Calorimetry Carcinoembryonic Antigen Cell Adhesion Cell Adhesion Molecules Cell Line Cells Chemicals Complementarity Determining Regions Cytoplasm Development Disease Drug Delivery Systems Electron Spin Resonance Spectroscopy Encapsulated Endocytosis Environment Enzyme Inhibitor Drugs Enzyme Inhibitors Escherichia coli Family Fibronectins Foundations Goals Health Height Hela Cells Heparin Human Integrins Life Lipid Bilayers Lipids Liposomes Maps Measures Mediating Membrane Membrane Proteins Meningitis Methods Micelles Molecular Mutagenesis N Domain Neisseria Neisseria gonorrhoeae Nuclear Magnetic Resonance Pathogenesis Pathway interactions Peptides Pharmacologic Substance Protein Binding Proteins Proteoglycan Relaxation Reporting Research Site Solutions Specificity Spectrum Analysis Spin Labels Structure Subgroup System Techniques Testing Therapeutic Thermodynamics Tissues Titrations Variant Vertebral column Work base design expectation extracellular insight interest mutant novel novel therapeutics protein structure protein structure function receptor reconstitution research study vaccine development

项目摘要

DESCRIPTION (provided by applicant): Many obligate bacterial membrane proteins hijack human cellular pathways by mimicking or manipulating host machinery. The goal of this research is to investigate the structure and dynamics of bacterial outer membrane proteins and their interactions with host receptors. Specifically, research is focused on the outer membrane opacity-associated proteins (Opa) from Neisseria gonorrhoeae and Neisseria meningitides, which induce engulfment of the bacterium in non-phagocytic cells by binding to host receptors. Opa proteins bind to various host receptors and are classified into two families based on host receptor selectivity. The larger class, OpaCEA, bind to carcinoembryonic antigen-like cellular adhesion molecules (CEACAMs), and the smaller class, OpaHS, bind to two different receptors; the heparansulfate proteoglycan receptors (HSPGs) directly and indirectly to integrin receptors via a heparin- mediated interaction with fibronectin or vibironectin. Opa proteins are integral outer membrane proteins and predicted to have an eight-stranded 2-barrel fold. Two of the extracellular loops (HV1 and HV2) have the most sequence variation between Opa proteins and determine the host receptors specificity. Not only do the HV loops discriminate between HSPG and CEACAM receptors, but OpaCEA proteins can be further divided into subgroups based on the selective binding to four of the seven CEACAM receptors. Using nuclear magnetic resonance, electron paramagnetic resonance, isothermal titration calorimetry, and mutagenesis, the molecular determinants of these interactions will be determined. The results will provide insight into the pathogenesis of Neisseria gonorrhoeae and Neisseria meningitides and, therefore, the potential for the rational design of novel antibiotics. In addition, the reconstituted Opa proteins may be useful for vaccine development. However, the most novel application of this research lies in the ability of Opa proteins to target host receptors specifically via three different mechanisms to induce endocytosis in non-phagocytic cells. This ability may be useful for liposome pharmaceutical carriers. The potential ability of liposome encapsulated therapeutics (e.g. enzymes, inhibitors, and peptides) to enter the cytoplasm of living cells and possibly tissue selectively is of crucial importance to the treatment of many diseases. Understanding the molecular determinants of the three Opa-mediated entry mechanisms may facilitate the development of liposome delivery mechanisms. PUBLIC HEALTH RELEVANCE: This research aims to determine how bacteria interact with human cells. By gaining an understanding of these molecular interactions, insights into the rational design of novel antibiotics, vaccine development, and targeting of liposome pharmaceutical carriers will be obtained.

描述（由申请人提供）：许多专性细菌膜蛋白通过模仿或操纵宿主机器来劫持人类细胞通路。这项研究的目的是研究细菌外膜蛋白的结构和动力学及其与宿主受体的相互作用。具体来说，研究重点是来自淋病奈瑟菌和脑膜炎奈瑟菌的外膜不透明相关蛋白（Opa），它们通过与宿主受体结合诱导非吞噬细胞中的细菌吞噬。 Opa 蛋白与各种宿主受体结合，并根据宿主受体选择性分为两个家族。较大的一类 OpaCEA 与癌胚抗原样细胞粘附分子 (CEACAM) 结合，较小的一类 OpaHS 与两种不同的受体结合；硫酸乙酰肝素蛋白聚糖受体（HSPG）通过肝素介导的与纤连蛋白或振动连蛋白的相互作用直接或间接地结合到整合素受体。 Opa 蛋白是完整的外膜蛋白，预计具有八链 2 桶折叠。两个细胞外环（HV1 和 HV2）在 Opa 蛋白之间具有最大的序列变异，并决定宿主受体的特异性。 HV 环不仅区分 HSPG 和 CEACAM 受体，而且 OpaCEA 蛋白还可以根据与七个 CEACAM 受体中的四个的选择性结合进一步分为亚组。使用核磁共振、电子顺磁共振、等温滴定量热法和诱变，将确定这些相互作用的分子决定因素。这些结果将有助于深入了解淋病奈瑟菌和脑膜炎奈瑟菌的发病机制，从而为合理设计新型抗生素提供潜力。此外，重组的 Opa 蛋白可能可用于疫苗开发。然而，这项研究最新颖的应用在于 Opa 蛋白能够通过三种不同的机制特异性靶向宿主受体，诱导非吞噬细胞的内吞作用。这种能力对于脂质体药物载体可能有用。脂质体封装的治疗剂（例如酶、抑制剂和肽）选择性地进入活细胞和可能的组织的细胞质的潜在能力对于许多疾病的治疗至关重要。了解三种 Opa 介导的进入机制的分子决定因素可能有助于脂质体递送机制的发展。公共健康相关性：这项研究旨在确定细菌如何与人体细胞相互作用。通过了解这些分子相互作用，将深入了解新型抗生素的合理设计、疫苗开发和脂质体药物载体的靶向。