Defining a novel mechanism of adhesion present in multiple infective endocarditis causing species

定义多种感染性心内膜炎引起物种中存在的新粘连机制

• 批准号: 10598825
• 负责人: Samantha Jane King
• 金额: $ 19.46万
• 依托单位: RESEARCH INST NATIONWIDE CHILDREN'S HOSP
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-12-05 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10598825
• 关键词: 3-Dimensional; Address; Adhesions; Affinity; Aging; Avidity; Bacteria; Bacterial Adhesins; Bacterial Adhesion; Binding; Binding Proteins; Blood Platelets; Carbohydrates; Cells; Complement; Congestive Heart Failure; Data; Development; Diagnostic; Endocardium; Family; Future; Health; Hospitalization; Human; Immunoglobulins; In Vitro; Infection; Infective endocarditis; Intervention; Knowledge; Lectin; Measures; Mediating; Microscopy; Mission; Modeling; Modification; Monitor; Morbidity - disease rate; Mutagenesis; Mutation; N-terminal; Orthologous Gene; Parents; Pathogenesis; Pathogenicity; Physiological; Polysaccharides; Population; Protein Family; Protein Region; Proteins; Public Health; Recombinant Proteins; Recombinants; Reporting; Research; Role; Serine; Sialic Acids; Specificity; Sterility; Streptococcus; Streptococcus Viridans Group; Streptococcus mitis; Streptococcus oralis; Structure; Surface; System; Testing; Therapeutic; Therapeutic Intervention; Time; United States National Institutes of Health; defined contribution; experimental study; inhibitor; innovation; member; mortality; mutant; novel; pathogen; polypeptide; prognostic; receptor; sialic acid binding Ig-like lectin; sialylation; targeted treatment; therapeutic target; three-dimensional modeling

The pathogenic mechanisms leading to subacute infective endocarditis (IE) are poorly understood, limiting the development of preventative and therapeutic strategies. However, bacterial binding to platelets is accepted as a key step in infection. Streptococcus oralis and Streptococcus mitis are viridans group streptococci that are common, but rarely studied, causes of IE. Terminal sialic acid is the major platelet receptor for many viridans group streptococci. Until recently, all known streptococcal-sialic acid interactions were mediated by members of the serine-rich repeat protein (SRRP) family of adhesins. That paradigm was shifted by identification of a novel sialic acid-binding protein, AsaA, in S. oralis. Binding of both SRRPs and AsaA is mediated by sialic acid immunoglobulin-like lectin (Siglec)-like domains. While most SRRPs contain one, AsaA contains two putative Siglec-like domains. The first Siglec-like domain contains a sialic acid-binding motif; however, the second Siglec-like domain is the first described to completely lack this motif. A polypeptide consisting of the two Siglec- like and Unique domains directly binds sialic acid on platelets. However, the role of each Siglec-like domain and the specific proteins bound are unknown. AsaA orthologs were identified in four IE-causing species with no previously described mechanisms of adhesion. Together, these data led to the hypothesis that AsaA and orthologs are members of a novel family of sialic acid-binding adhesins, that contribute to IE by binding host components via two Siglec-like domains. Three aims will test this hypothesis. Aim 1: Determine the role of the two AsaA Siglec-like domains in binding IE-relevant host components. Defined mutants and recombinantly expressed proteins will be used to resolve the contribution of the two different Siglec-like domains to platelet binding and to identify the specific proteins bound by each domain. Furthermore, the range of IE-relevant host components bound by AsaA and the role of the two Siglec-like domains in these interactions will be established. Aim 2: Determine whether AsaA is a member of a novel family of sialic acid-binding adhesins. The role of AsaA orthologs in platelet binding will be investigated, with a focus on S. mitis. Aim 3: Determine the contribution of AsaA, and specifically sialic acid binding, to IE using a 3D human microvessel model. A cutting- edge physiologically relevant and tractable 3D microvessel model will be used to assess the role of sialic acid, AsaA, and the Siglec-like domains in establishment of a vegetation. Furthermore, experiments will ascertain whether this adhesion mechanism could be a therapeutic target. This application is significant as it focuses on bacteria with no previously described mechanisms of binding platelets, a critical step in development of subacute IE. Innovative aspects of this proposal include defining the function of a novel Siglec-like domain and utilizing a cutting-edge 3D microvessel model. This study will also likely identify a novel family of sialic acid- binding adhesins produced by multiple IE-causing species, which may bind the same receptor as SRRPs making these interactions an attractive target for intervention.

导致亚急性感染性心内膜炎（IE）的致病机制知之甚少，限制了 制定预防和治疗策略。然而，细菌与血小板的结合被认为是 感染的关键一步。口腔链球菌和轻链球菌是草绿色链球菌，它们是 IE 的常见但很少研究的原因。末端唾液酸是许多草绿色的主要血小板受体 链球菌群。直到最近，所有已知的链球菌-唾液酸相互作用都是由成员介导的 富含丝氨酸重复蛋白（SRRP）的粘附素家族。通过确定一个 口腔链球菌中新型唾液酸结合蛋白 AsaA。 SRRP 和 AsaA 的结合均由唾液酸介导 免疫球蛋白样凝集素 (Siglec) 样结构域。虽然大多数 SRRP 包含一个，但 AsaA 包含两个假定的 类似 Siglec 的域。第一个 Siglec 样结构域包含唾液酸结合基序；然而，第二个 Siglec 样结构域是第一个被描述为完全缺乏该基序的结构域。由两个Siglec-组成的多肽 like 和 Unique 结构域直接结合血小板上的唾液酸。然而，每个 Siglec 类域的作用 并且结合的具体蛋白质是未知的。在四种引起 IE 的物种中发现了 AsaA 直向同源物，但没有发现 先前描述的粘附机制。这些数据共同得出这样的假设：AsaA 和 直向同源物是唾液酸结合粘附素新家族的成员，通过结合宿主来促进 IE 通过两个类似 Siglec 的域组成。三个目标将检验这一假设。目标 1：确定角色 两个 AsaA Siglec 样域用于绑定 IE 相关的宿主组件。确定的突变体和重组体 表达的蛋白质将用于解析两个不同的 Siglec 样结构域对血小板的贡献 结合并识别每个结构域结合的特定蛋白质。此外，IE相关主机的范围 由 AsaA 结合的成分以及两个 Siglec 样结构域在这些相互作用中的作用将是 已确立的。目标 2：确定 AsaA 是否是唾液酸结合粘附素新家族的成员。这 将研究 AsaA 直向同源物在血小板结合中的作用，重点是轻链球菌。目标 3：确定 使用 3D 人体微血管模型研究 AsaA（特别是唾液酸结合）对 IE 的贡献。一个切割—— 边缘生理相关且易于处理的 3D 微血管模型将用于评估唾液酸的作用， AsaA 和 Siglec 类域在植被建立中的作用。此外，实验将确定 这种粘附机制是否可以成为治疗靶点。该应用程序很重要，因为它重点关注 以前没有描述过的结合血小板机制的细菌，这是开发中的关键一步 亚急性IE。该提案的创新方面包括定义新的 Siglec 类域的功能和 利用尖端的 3D 微血管模型。这项研究还可能鉴定出一个新的唾液酸家族—— 多种引起 IE 的物种产生的结合粘附素，可能与 SRRP 结合相同的受体 使这些相互作用成为有吸引力的干预目标。