Structure and dynamics of bacterial membrane protein - receptor interactions

描述(申请人提供)：许多专性细菌膜蛋白通过模仿或操纵宿主机制劫持人类细胞途径。本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。具体地说，研究重点是淋球菌和脑膜炎奈瑟菌的外膜不透明相关蛋白(OpA)，它通过与宿主受体结合来诱导细菌在非吞噬细胞中被吞噬。OPA蛋白与不同的宿主受体结合，根据宿主受体的选择性分为两个家族。较大的OpaCEA与癌胚抗原样细胞黏附分子(CEACAM)结合，较小的OPAHS与两种不同的受体结合；肝素蛋白多糖受体(HSPGs)通过肝素介导的与纤维连接蛋白或葡萄连蛋白的相互作用直接或间接与整合素受体结合。OPA蛋白是一种完整的外膜蛋白，预测有一个8链2桶折叠。其中两个胞外环(Hv1和Hv2)在OpA蛋白之间具有最大的序列差异，并决定了宿主受体的特异性。HV环不仅区分HSPG和CEACAM受体，而且OpaCEA蛋白还可以根据与七种CEACAM受体中的四种的选择性结合而进一步划分为亚群。利用核磁共振、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。这一结果将为淋球菌和脑膜炎奈瑟菌的发病机制提供洞察力，从而为合理设计新型抗生素提供可能性。此外，重组的OpA蛋白可能对疫苗开发有用。然而，这项研究最新的应用在于，OpA蛋白能够通过三种不同的机制特异性地靶向宿主受体，诱导非吞噬细胞的内吞作用。这种能力可能对脂质体药物载体有用。脂质体包裹的治疗药物(如酶、抑制剂和多肽)选择性地进入活细胞和组织的细胞质的潜在能力对许多疾病的治疗至关重要。了解OpA介导的三种进入机制的分子决定因素可能有助于脂质体给药机制的发展。

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.