Inhibition of the Classical & Lectin Complement Pathways by Staphylococcus aureus Eap

古典的抑制

• 批准号: 8891551
• 负责人: Brian V Geisbrecht
• 金额: $ 18.75万
• 依托单位: KANSAS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-01-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8891551
• 关键词: Active Sites; Acute; Adherence; Affinity; Anti-Inflammatory Agents; Anti-inflammatory; Autoimmune Diseases; Bacteria; Basic Science; Binding; Binding Sites; Biochemical; Biological Assay; Cell surface; Chronic; Complement; Complement 3 Convertase; Complement 3b; Complement 4b; Complement Activation; Complement Inactivators; Complex; Cyclic Peptides; Development; Disease; Dose; Enzymes; Event; Extracellular Domain; Extracellular Protein; FDA approved; Foundations; Future; Generations; Goals; Graft Rejection; Human; Human body; Hyperactive behavior; Immunity; Inflammation; Inflammation Mediators; Inflammatory; Intervention; Investigation; Lead; Lectin; Libraries; Malignant Neoplasms; Mediating; Modeling; Molecular; Mutagenesis; Nature; Neurodegenerative Disorders; Pathway interactions; Peptide Hydrolases; Peptides; Phage Display; Pharmaceutical Preparations; Play; Price; Process; Protein Binding; Protein Family; Protein Inhibition; Proteins; Rare Diseases; Regulation; Reperfusion Injury; Role; Route; Sepsis Syndrome; Series; Serine Protease; Serpins; Site; Specificity; Staphylococcus aureus; Structure; Structure-Activity Relationship; System; Therapeutic; Virulent; Whole Blood; Work; alternative pathway complement C3 convertase; base; biomaterial incompatibility; complement 4b-binding protein; complement C2a; complement C3 precursor; complement C4c; complement pathway; complement system; design; driving force; extracellular; human disease; inhibitor/antagonist; insight; member; neutrophil; novel; novel strategies; pathogen; protein structure function; public health relevance; response; screening; therapeutic target

 DESCRIPTION (provided by applicant): Over the last several years, our understanding of the complement evasion mechanisms utilized by pathogens has increased precipitously through the study of the virulent bacterium Staphylococcus aureus. By screening a library of secreted S. aureus proteins in a human whole-blood model of inflammation, we have identified the Extracellular Adherence Protein (Eap) as the first known S. aureus inhibitor of the Classical (CP) and Lectin (LP) pathways of complement. Eap inhibits both of these pathways in a dose-dependent manner that requires formation of high-nanomolar affinity interaction with complement component C4b. This interaction blocks formation of the CP/LP pro-C3 convertase complex (C4b/C2), which dramatically lowers levels of the active CP/LP C3 convertase (C4b/C2a). Using the same whole-blood model, we have also identified Eap as a potent inhibitor of Neutrophil Serine Proteases (NSPs). Unlike conventional serpins, Eap inhibition of NSPs is non-covalent in nature. Furthermore, it occurs through a molecular mechanism distinct from its effects on the complement system since two related proteins, EapH1 and EapH2, also block NSP activity but have no effect on complement. In this proposal, we will investigate the molecular basis for the specificity of Eap's effects on the CP/LP through two Specific Aims: (1) We will characterize the biochemical and structural basis for Eap binding to complement protein C4b, and (2) We will characterize peptides that compete with Eap for C4b binding and determine whether they retain Eap-like inhibitory activities against the CP/LP. We expect that this integrated structure/function and discovery approach will provide new insight into regulation of the CP/LP. In turn, this may hold important clues into the design and optimization of novel complement-targeted, anti-inflammatory therapeutics in the future.

 描述（由申请方提供）：在过去几年中，通过对强毒细菌金黄色葡萄球菌的研究，我们对病原体利用的补体逃避机制的理解急剧增加。通过筛选分泌型S.金黄色葡萄球菌蛋白在人全血炎症模型中的表达，我们已经鉴定了细胞外粘附蛋白（Eap）作为第一个已知的金黄色葡萄球菌。金黄色葡萄球菌补体经典（CP）和凝集素（LP）途径的抑制剂。Eap以剂量依赖性方式抑制这两种途径，需要与补体成分C4 b形成高纳摩尔亲和力相互作用。这种相互作用阻断CP/LP pro-C3转化酶复合物（C4 b/C2）的形成，这显著降低了活性CP/LP C3转化酶（C4 b/C2 a）的水平。使用相同的全血模型，我们还将Eap鉴定为神经元丝氨酸蛋白酶（NSP）的有效抑制剂。与常规丝氨酸蛋白酶抑制剂不同，NSP的Eap抑制本质上是非共价的。此外，它通过与其对补体系统的作用不同的分子机制发生，因为两种相关蛋白EapH 1和EapH 2也阻断NSP活性，但对补体没有影响。在这个提议中，我们将通过两个特定的目的来研究Eap对CP/LP作用的特异性的分子基础：（1）我们将表征Eap与补体蛋白C4 b结合的生物化学和结构基础，以及（2）我们将表征与Eap竞争C4 b结合的肽，并确定它们是否保留对CP/LP的Eap样抑制活性。我们期望这种整合的结构/功能和发现方法将为CP/LP的调控提供新的见解。反过来，这可能为未来新型补体靶向抗炎治疗的设计和优化提供重要线索。