Investigation of FadA adhesin from Fusobacterium nucleatum

具核梭杆菌 FadA 粘附素的研究

• 批准号: 7736929
• 负责人: Yiping Han
• 金额: $ 35.05万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7736929
• 关键词: Abscess; Address; Affect; Amino Acids; Anaerobic Bacteria; Animal Model; Apoptosis; Attention; Award; Bacteria; Bacterial Adhesins; Binding; Binding Sites; Biological Process; Body part; Cell Adhesion; Cell Adhesion Molecules; Cell Communication; Cell physiology; Cell-Cell Adhesion; Cells; Complex; Diagnosis; Discipline of obstetrics; Disease; Drug Delivery Systems; Endothelial Cells; Epithelial; Filament; Focal Adhesions; Foundations; Fusobacteria; Fusobacterium; Fusobacterium nucleatum; Goals; Growth; Head; Hematogenous; Immune response; In Vitro; Incidence; Infection; Inflammatory Response; Invaded; Investigation; Leucine; Light; Mediating; Membrane; Membrane Proteins; Modeling; Mus; Neoplasm Metastasis; Oral; Oral cavity; Organism; Pathogenesis; Pathway interactions; Pattern; Peptide Signal Sequences; Peptides; Periodontal Diseases; Placenta; Play; Pregnancy Outcome; Premature Birth; Proteins; Receptor Cell; Research; Resolution; Role; Screening procedure; Site; Structure; Structure-Activity Relationship; System; Tail; Testing; Tissues; Uncertainty; Virulent; Wound Healing; Yeasts; alpha helix; angiogenesis; biological adaptation to stress; cancer cell; cell motility; frontier; in vitro testing; in vivo; innovation; intraamniotic infection; microorganism; monomer; mutant; novel; oral bacteria; oral biology; pathogen; pregnant; public health relevance; receptor; receptor binding; therapeutic development; therapeutic target; tissue culture; transmission process; yeast two hybrid system

DESCRIPTION (provided by applicant): Fusobacterium nucleatum is a Gram-negative anaerobe implicated in various forms of periodontal diseases. It is also associated with infections in other parts of the body and is one of the most prevalent species in intra- amniotic infection, causing preterm birth. F. nucleatum binds to and invades host epithelial and endothelial cells, a mechanism allowing colonization at different host sites. Previous studies have shown that F. nucleatum can translocate to the pregnant mouse placenta via haematogenous transmission, followed by activation of localized placental inflammatory responses, leading to adverse pregnancy outcomes. Invasion of mouse placental endothelial cells by F. nucleatum has also been observed in vivo. So far, only one adhesin, FadA (for Fusobacterium adhesin A), has been identified to be required for bacterial binding and invasion of host cells in both tissue-culture and animal models. FadA is a unique adhesin consisting of two forms: the intact non-secreted form (pre-FadA) composed of 129 amino-acid (aa) residues, and the mature secreted form (mFadA) of 111 aa. The crystal structure of mFadA reveals two anti-parallel alpha-helices connected by an 8- aa loop. The crystal structure of mFadA suggests oligomerization in a head-to-tail pattern via a novel "leucine chain" motif. Filament formation and binding to host cells require both pre-FadA and mFadA. A FadA adhesin model has been proposed, with pre-FadA anchored in the inner membrane and a chain of mFadA on top of pre-FadA protruding through the outer membrane. We hypothesize that the receptor-binding site may be located in the loop region, only fully exposed at the tip of the filament. Furthermore, several of the FadA filaments may bundle together to form a cluster of loops, which may be required for binding. One focus of this proposed study is to test the FadA adhesin model. Using a yeast-two-hybrid system, several putative receptors have been identified to interact with FadA. Thus, a second focus of this study is to validate the interactions between the putative receptors and FadA and to investigate the host responses to FadA. Our specific aims are: Aim I. Further characterization of the FadA adhesin in F. nucleatum. Five sub-aims are proposed: (i) investigating fadA expression under different conditions, (ii) investigating the spatial arrangement of FadA in F. nucleatum, (iii) investigating the involvement of the loop region in host-cell binding, (iv) investigating the role of the signal peptide in FadA complex formation, and (v) investigating possible accessory molecules in F. nucleatum associated with FadA. Aim II. Investigation of FadA and host cell interactions. Two sub-aims are proposed: (i) continued identification and characterization of the FadA receptor, and (ii) investigating the effect of FadA on cellular processes. From this study, we hope to identify (i) potential therapeutic targets for inhibiting F. nucleatum colonization in the host, and (ii) host components and pathways affected by FadA which will facilitate modulation of respective cellular processes and targeted drug delivery. PUBLIC HEALTH RELEVANCE: In this competitive renewal application, we propose to continue in-depth investigation of the novel FadA adhesin from oral bacterium Fusobacterium nucleatum, which is associated with periodontal disease and preterm birth. FadA plays an important role in the bacterial colonization in the host and in causing preterm birth. Continued structure-function analysis of FadA will facilitate therapeutic development to inhibit the bacterial colonization in the host and to reduce the incidence of preterm birth. Identification of host components and pathways affected by FadA will facilitate therapeutic development of modulate specific cellular processes and targeted drug delivery.

描述（由申请人提供）：fusobacterium nucleatum是一种革兰氏阴性厌食症，涉及各种形式的牙周疾病。它也与体内其他部位的感染有关，是羊膜内感染中最普遍的物种之一，导致早产。 F.核与宿主上皮和内皮细胞结合并入侵，这是一种允许在不同宿主部位定殖的机制。先前的研究表明，F. nucleatum可以通过血液传播转移到孕妇小鼠胎盘，然后激活局部胎盘炎症反应，从而导致不良妊娠结局。还观察到在体内观察到小鼠胎盘内皮细胞的侵袭。到目前为止，只有一种粘附素FADA（用于粘杆菌A）已被确定为在组织培养和动物模型中宿主细胞的细菌结合和侵袭所必需的。 FADA是一种独特的粘附蛋白，由两种形式组成：完整的未分泌形式（PreFADA）由129个氨基酸（AA）残基组成，而成熟的分泌形式（MFADA）为111 aa。 MFADA的晶体结构揭示了通过8 aa环连接的两个反行α-螺旋。 MFADA的晶体结构表明，通过新颖的“亮氨酸链”基序以头到尾模式中的寡聚化。细丝形成和与宿主细胞的结合需要前法和MFADA。已经提出了FADA粘附模型，前fada锚定在内膜中，并在通过外膜突出的前fada链上。我们假设受体结合位点可能位于环区域，只有完全暴露在细丝的尖端。此外，一些FADA丝可能会捆绑在一起以形成一个循环，这可能是绑定所必需的。这项拟议的研究的一个重点是测试FADA粘附素模型。使用酵母 - 两种杂交系统，已经确定了几种推定的受体与FADA相互作用。因此，这项研究的第二个重点是验证推定受体与FADA之间的相互作用，并研究宿主对FADA的反应。我们的具体目的是：目标I。FADA粘附蛋白在F. nucleatum中的进一步表征。提出了五个子-IAM：（i）研究FADA在不同条件下的表达，（ii）研究FADA在F. nucleatum中的空间排列，（iii）研究环路区域在宿主细胞结合中的参与，（iv）研究信号肽在FADA复合物中的作用，（V）与FADA复合物的作用相关的FADA综合物，以及（V）可能与FACE的访问相关化的成分。目标II。研究FADA和宿主细胞相互作用。提出了两个子-IAM：（i）FADA受体的持续鉴定和表征，以及（ii）研究FADA对细胞过程的影响。从这项研究中，我们希望确定（i）潜在的治疗靶标，以抑制宿主中的核核定植，以及（ii）受FADA影响的宿主成分和途径，这将有助于调节各自的细胞过程和靶向药物的递送。 公共卫生相关性：在这种具有竞争性的更新应用中，我们建议继续深入研究来自口腔细菌核细菌核细菌的新型FADA粘附素，这与牙周疾病和早产有关。 FADA在宿主的细菌定殖并导致早产中起重要作用。 FADA的持续结构功能分析将促进治疗发育，以抑制宿主的细菌定植并减少早产的发生率。鉴定受FADA影响的宿主成分和途径将促进调节特定细胞过程和靶向药物递送的治疗发展。