Dissecting the role of sialic acid and sialidase in the pathophysiology of Porphyromonas gingivalis

剖析唾液酸和唾液酸酶在牙龈卟啉单胞菌病理生理学中的作用

• 批准号: 10545715
• 负责人: Chunhao Chris Li
• 金额: $ 48.92万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-15 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10545715
• 关键词: 3-Dimensional; Address; Adherence; Alveolar Bone Loss; Alzheimer' s Disease; Anabolism; Arthritis; Bacteria; Binding; Binding Sites; Biochemical; Biochemistry; Biological; Biological Assay; Biological Markers; Biological Process; Biophysics; Carbon; Cardiovascular Diseases; Cell Communication; Cell Surface Receptors; Cell secretion; Cell surface; Cells; Charge; Clinical Research; Complement; Crystallography; Data; Development; Diabetes Mellitus; Diagnosis; Dose; Embryonic Development; Enzymes; Excision; Forsythia; Functional disorder; Future; Genes; Genetic; Genome; Gingival Crevicular Fluid; Glycobiology; Glycoconjugates; Goals; Growth; Homologous Gene; Human; Immune; Immune Evasion; Immune system; Immunoglobulins; Immunology; Impairment; In Vitro; Innate Immune Response; Intercept; Invaded; Kinetics; Ligands; Lipopolysaccharides; Macrophage; Maps; Measures; Mediating; Membrane Proteins; Microbial Biofilms; Modality; Molecular; Mucins; Mucous Membrane; Neuraminic Acids; Neuraminidase; Neurologic; Nutrient; Oral cavity; Osteitis; Pathogenesis; Pathogenicity; Pathway interactions; Periodontal Diseases; Periodontitis; Phagocytosis; Phenotype; Play; Polysaccharides; Porphyromonas gingivalis; Positioning Attribute; Proteins; Recommendation; Reporting; Resistance; Role; Salivary; Series; Severities; Shapes; Sialic Acids; Side; Structure; Substrate Specificity; Sugar Acids; Surface; Testing; Toll-like receptors; Vagina; Virulence; Virulence Factors; antimicrobial; biological adaptation to stress; capsule; cell growth; chemokine; complement system; design; dietary; experimental study; fitness; genetic approach; gut bacteria; gut microbiome; immune function; immunoregulation; in vivo; inhibitor; insight; interdisciplinary approach; mouse model; nano; neutrophil; novel; oral bacteria; oral pathogen; pathogen; pathogenic bacteria; patient response; polymicrobial biofilm; prevent; response; sialic acid permease; sialylation; structural biology; treatment response; uptake

The overarching goal of this application is to investigate the role of sialidase and sialic acid in the keystone pathogen Porphyromonas gingivalis (Pg) and their contributions to the pathogenesis of periodontitis. Sialic acid (SA), a group of structurally related nine-carbon sugar acids, plays critical roles in host-pathogen interactions. On the host side, mammalian mucosal surfaces and secretions of the mouth, airway, gut, and vagina are especially sialoglycan-rich, which have a variety of biological, biophysical, antimicrobial, and immunological functions. In addition, a number of cell surface receptors (e.g., chemokine-, immunoglobulin-, and toll-like- receptors) are either sialylated or recognize sialylated ligands, which play critical roles in immune recognition and activation. On the pathogen side, many bacterial pathogens have evolved different mechanisms to target host SA for adherence, invasion, immune modulation and nutrient acquisition, thereby promoting their fitness and pathogenesis. Specifically, bacterial pathogens often use sialidases to hydrolyze host sialoglycans, compromise host immune defenses, and promote their survival in the mucosal niche. Salivary and gingival crevicular fluids contain a high concentration of SA bound to sialoglycans in various proteins such as mucins. Clinical studies indicate that sialidase activity in the oral cavity is positively associated with the severity of periodontitis; thus, it is recommended as a biomarker for periodontitis diagnosis. Accordingly, sialidases have been found in numerous oral bacteria including the keystone pathogen Porphyromonas gingivalis (Pg). Pg lacks genes to synthesize SA. Instead, it encodes a sialidase (PG0352), which is highly conserved among all genome sequenced Pg isolates. Previous studies from our group and others have shown that PG0352 plays a crucial role in Pg capsule synthesis, biofilms, stress response, innate immune responses, and virulence. However, the molecular mechanism underlying these phenotypes remains elusive. In this application, we hypothesize that Pg employs a sialidase and a SA specific transporter to scavenge host SA, which is in turn used to modify Pg cell surface molecules such as capsule and lipopolysaccharide (LPS), thereby intercepting host innate immune defenses, such as complement killing and phagocytosis of neutrophils and macrophage. To test this hypothesis, the following studies are specifically designed and will be implemented: Aim 1: To delineate the biochemical and structural features of PG0352 by using an approach of genetics, biochemistry, and crystallography; Aim 2: To investigate how Pg imports and utilizes SA by using a multidisciplinary approach of genetics, biochemistry, glycobiology, immunology, and biofilm assays; and Aim 3: To elucidate the role of PG0352 in the pathogenicity of Pg by using various in vitro and in vivo approaches. Completion of this project will not only provide mechanistic insights into understanding the role of SA and sialidase in the pathophysiology of Pg and perhaps other oral pathogens as well, but also will pave a way for future development of specific sialidase inhibitors against oral pathogens, which can provide alternatives to mitigate periodontal diseases.

该应用的总体目标是研究唾液酸酶和唾液酸在基石中的作用 病原体卟啉单胞菌（PG）及其对牙周炎发病机理的贡献。唾液酸 （SA）是一组与结构相关的九个碳糖酸，在宿主 - 病原体相互作用中起着关键作用。 在宿主一侧，哺乳动物的粘膜表面和嘴，气道，肠道和阴道的分泌物是 特别是唾液酸富集，具有多种生物学，生物物理，抗菌和免疫学 功能。另外，许多细胞表面受体（例如趋化因子，免疫球蛋白和Toll-like- 受体）是脱糖化的或识别脱脂的配体，在免疫识别中起关键作用 和激活。在病原体方面，许多细菌病原体已经进化了不同的机制 主持人SA依从性，入侵，免疫调节和养分获取，从而促进其健康状况 和发病机理。具体而言，细菌病原体经常使用唾液酸酶水解宿主的唾液聚糖， 妥协宿主的免疫防御，并促进其在粘膜细分市场中的生存。唾液和牙龈 缝隙液在各种蛋白质（如粘蛋白）中含有高浓度与唾液聚糖结合的SA。 临床研究表明，口腔中的唾液酸酶活性与 牙周炎；因此，建议作为牙周炎诊断的生物标志物。因此，唾液酸酶具有 在许多口腔细菌中发现了包括基石病原体卟啉单胞菌（PG）。 pg 缺乏基因来合成SA。取而代 基因组测序的PG分离株。我们小组和其他人的先前研究表明，PG0352扮演 在PG胶囊合成，生物膜，压力反应，先天免疫反应和毒力中的关键作用。 然而，这些表型基础的分子机制仍然难以捉摸。在此应用程序中，我们 假设PG使用唾液酸酶和SA特异性转运蛋白来清除宿主SA，而这又是 用于修改PG细胞表面分子，例如胶囊和脂多糖（LPS），从而拦截 宿主先天免疫防御，例如中性粒细胞和巨噬细胞的补体杀伤和吞噬作用。到 检验此假设，以下研究是专门设计的，并且将被实施：目标1：描述 PG0352的生化和结构特征通过使用遗传学，生化和 晶体学；目标2：调查PG如何通过使用多学科方法进口SA并使用SA 遗传学，生物化学，糖生物学，免疫学和生物膜测定；目标3：阐明 PG0352通过使用各种体外和体内方法在PG的致病性中。该项目的完成 不仅将提供理解SA和唾液酸酶在病理生理学中的作用的机械见解 PG，也许还有其他口腔病原体，但也将为未来开发特定的方式铺平 唾液酸酶抑制剂针对口腔病原体，可以提供减轻牙周疾病的替代方法。