Mechanisms of Pneumococcal Adherence

肺炎球菌粘附机制

• 批准号: 8096644
• 负责人: Samantha Jane King
• 金额: $ 31.76万
• 依托单位: RESEARCH INST NATIONWIDE CHILDREN'S HOSP
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8096644
• 关键词: Adherence; Antibiotic Resistance; Bacteria; Bacterial Adhesins; Binding; Binding Sites; Biochemical; C-terminal; Cell surface; Cessation of life; Clinical; Complement; Conflict (Psychology); Conjugate Vaccines; Data; Development; Disease; Enzymes; Epithelial; Epithelial Cells; Epithelial Receptor Cell; Exoglycosidases; Galactosidase; Glycoside Hydrolases; Goals; Health; Human; Knowledge; Ligands; Lipids; Measures; Mediating; Molecular; Mutagenesis; N-terminal; Neuraminidase; Otitis Media; Pathogenesis; Pneumococcal Infections; Polysaccharides; Preventive; Protein Region; Proteins; Recombinants; Reporting; Respiratory System; Respiratory tract structure; Role; Scanning; Serotyping; Site-Directed Mutagenesis; Streptococcus pneumoniae; Structure; Surface; Techniques; Therapeutic; Vaccines; X-Ray Crystallography; airway epithelium; bacterial adhesin receptor; capsule; in vivo; innovation; insight; novel; pathogen; prevent; receptor

DESCRIPTION (provided by applicant): S. pneumoniae (S.p.) is an important human pathogen causing more than 1 million deaths per year world-wide. In addition, S.p is responsible for an estimated 7 million cases of otitis media each year in the US alone. The currently available conjugate vaccine includes 7 of the 90 known capsule types. While this vaccine has significantly reduced invasive disease and colonization caused by vaccine serotypes, there has been an unexpected increase in both invasive disease and colonization caused by non-vaccine serotypes. In addition, the vaccine does not effectively protect against otitis media. Colonization of the respiratory tract is an essential precursor to disease. Despite the importance of adherence to colonization and, therefore, pathogenesis, the mechanisms by which S.p adheres to airway epithelia remain unclear. We have identified a novel exoglycosidase-dependent adherence mechanism requiring the S.p. neuraminidase, NanA, and 2- galactosidase, BgaA. This mechanism is relevant to adherence of recent clinical isolates and S.p. adherence to human primary epithelial cells, suggesting that it will be relevant in vivo. Our preliminary data demonstrate that while NanA exposes a receptor for adherence, BgaA acts as an adhesin. To further define this mechanism of adherence we have 2 Specific Aims: 1) Elucidate the BgaA binding site. We will define the BgaA region that mediates adherence to human epithelial cells. BgaA is the 2nd largest protein expressed by S.p. and possesses a C-terminal region of unknown function. We will perform molecular and structural analyses to elucidate how BgaA acts as an adhesin. Different regions of the protein will be expressed and used in binding and inhibition studies to identify the region(s) that mediate(s) adherence. The residues that contribute to binding will subsequently be identified by mutagenesis scanning. Structuring of BgaA will be utilized as an alternative approach to identify the binding site and identify the function of the C-terminal region of the protein. 2) Identify the BgaA receptor on the epithelial cell surface. The initial attachment of S.p. has been proposed to occur through a glycan receptor exposed by neuraminidase; however, the adhesin is unknown and there have been conflicting reports of different receptors. We will use a comprehensive range of techniques to determine if BgaA binds to a glycan and if this is structure in the context of a protein or lipid. Following identification of the receptor we will further characterize its interaction with BgaA. The successful completion of these Specific Aims will provide an increased understanding of this mechanism of S.p. adherence. This knowledge is likely to contribute significantly to the long-term goal of developing more effective vaccines and/or treatments to reduce the burden of S.p. disease. PUBLIC HEALTH RELEVANCE: Colonization of the human airway is an essential precursor to pneumococcal disease; however the mechanisms by which the bacteria initially attach to the airway are poorly understood. This study will characterize a novel glycosidase-dependent mechanism of adherence. We will identify the bacterial adhesin and the receptor on the epithelial surface. Our increased understanding of pneumococcal adherence will help us achieve the longer term goal of developing a more effective vaccine or treatment to reduce the burden of pneumococcal disease.

描述（由申请人提供）：肺炎链球菌 (S.p.) 是一种重要的人类病原体，每年在全世界造成超过 100 万人死亡。此外，仅在美国，每年估计就有 700 万例中耳炎病例由 S.p 引起。目前可用的结合疫苗包括 90 种已知胶囊类型中的 7 种。虽然这种疫苗显着减少了由疫苗血清型引起的侵袭性疾病和定植，但由非疫苗血清型引起的侵袭性疾病和定植却意外增加。此外，该疫苗不能有效预防中耳炎。呼吸道定植是疾病的重要先兆。尽管粘附定植及其发病机制很重要，但 S.p 粘附气道上皮的机制仍不清楚。我们已经确定了一种新的外切糖苷酶依赖性粘附机制，需要 S.p.神经氨酸酶、NanA 和 2-半乳糖苷酶、BgaA。该机制与最近的临床分离株和 S.p. 的依从性有关。与人原代上皮细胞的粘附，表明它在体内具有相关性。我们的初步数据表明，虽然 NanA 暴露了粘附受体，但 BgaA 充当粘附素。为了进一步定义这种粘附机制，我们有 2 个具体目标：1) 阐明 BgaA 结合位点。我们将定义介导人上皮细胞粘附的 BgaA 区域。 BgaA 是 S.p. 表达的第二大蛋白质。并具有未知功能的C端区域。我们将进行分子和结构分析，以阐明 BgaA 如何充当粘附素。蛋白质的不同区域将被表达并用于结合和抑制研究，以鉴定介导粘附的区域。随后将通过诱变扫描来鉴定有助于结合的残基。 BgaA 的构建将用作识别结合位点和识别蛋白质 C 末端区域功能的替代方法。 2) 识别上皮细胞表面的BgaA受体。 S.p. 的初始附件。已提出通过神经氨酸酶暴露的聚糖受体发生；然而，粘附素是未知的，并且不同受体的报道相互矛盾。我们将使用一系列全面的技术来确定 BgaA 是否与聚糖结合，以及这是否是蛋白质或脂质中的结构。在鉴定出受体后，我们将进一步表征其与 BgaA 的相互作用。这些具体目标的成功完成将加深对 S.p. 机制的理解。坚持。这些知识可能会对开发更有效的疫苗和/或治疗方法以减轻链球菌负担的长期目标做出重大贡献。疾病。公共卫生相关性：人类气道定植是肺炎球菌疾病的重要先兆；然而，人们对细菌最初附着在气道上的机制知之甚少。这项研究将描述一种新的糖苷酶依赖性依从机制。我们将识别细菌粘附素和上皮表面上的受体。我们对肺炎球菌依从性的更多了解将有助于我们实现开发更有效的疫苗或治疗方法以减轻肺炎球菌疾病负担的长期目标。