Functional Domains of Borrelia burgdorferi Virulence Factor OspC

伯氏疏螺旋体毒力因子 OspC 的功能域

• 批准号: 8772760
• 负责人: WOLFRAM R ZUECKERT
• 金额: $ 18.88万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8772760
• 关键词: Architecture; Bacteria; Binding; Biological Assay; Biological Process; Borrelia; Borrelia burgdorferi; Borrelia turicatae; Cell surface; Chimera organism; Cysteine; Disease; Distal; Event; Family; Gram' s stain; Gram-Negative Bacteria; Human; Immune; In Vitro; Infection; Intervention; Investigation; Lead; Length; Lipids; Lipoproteins; Lyme Disease; Mediating; Membrane; Mus; N-terminal; Nutrient; Order Spirochaetales; OspC protein; Peptides; Positioning Attribute; Prevalence; Property; Proteins; Proteome; Publishing; Research; Role; Serotyping; Sorting - Cell Movement; Structure; Structure-Activity Relationship; Surface; Testing; Ticks; Translating; United States; Vaccines; Vector-transmitted infectious disease; Virulence Factors; Work; design; feeding; mouse model; mutant; novel; outer surface lipoprotein; pathogen; protein folding; public health relevance; relapsing fever borrelia; routine Bacterial stain; scaffold; three dimensional structure; tissue tropism; vector

DESCRIPTION (provided by applicant): Lyme disease, the most common vector-borne disease in the Northern hemisphere, is transmitted to humans by ticks carrying the spirochete Borrelia burgdorferi. Enabled by a sudden influx of nutrients during tick feeding, the bacterium quickly remodels its surface to allow for efficient establishment of mammalian infection. The B. burgdorferi host-pathogen interface is dominated by lipoproteins that are peripherally anchored in the outer leaflet of the outer membrane via a lipid-modified cysteine. Recent studies in a mouse model of infection have shown that expression of outer surface lipoprotein C (OspC) is crucial for establishment of early infection. The overall objective of this study is to determine te structure function relationships of this important spirochetal virulence factor. OspC forms a homodimer on the surface of the cells, and its three-dimensional structure can be separated into three distinct domains: (i) a disordered N-terminal "tether" peptide, (ii) an alpha helical bundle "scaffold", and (iii) a membrane-distal "dome" of helix-connecting loops. Yet, it remains unclear how these three domains are contributing to the lipoprotein's biological function in bacterial colonization and dissemination. OspC belongs to the same protein fold family as the relapsing fever Borrelia Vsp proteins, which are involved in immune evasion and tissue tropism. In fact, the crystal structure of Borrelia turicatae Vsp1 deviates from OspC only slightly but most significantly in the helix-connecting loops. We therefore hypothesize that OspC's dome residues are crucial for OspC's specific function during early infection. Furthermore, we hypothesize that the tether peptide, in addition to providing protein sorting information (see our published work), evolved to properly position the virulence factor within the bacterium's surface proteome through optimization of its length. We will test these hypotheses in the following two specific aims: 1. T determine the role of OspC dome residues in colonization and dissemination by testing if Vsp proteins or Vsp scaffold-OspC dome chimeras can functionally replace OspC both in binding of host molecules in vitro as well as in in a mouse model of infection. Ultimately, the minimal OspC dome domain required for function will be identified. 2. To determine the role of OspC lipoprotein tether length in virulence factor accessibility and function by testing existing and newly generated OspC tether length mutants for functions in vitro as well as in a competitive mouse infection assay. These studies will (i) achieve further milestones in our investigation of spirochetal surface lipoprotein structure- function, (ii) significantly increase our understanding f the early events during mammalian infection with Borrelia burgdorferi, and (iii) yield important clues that may ultimately translate into the design of novel intervention strategies for Lyme disease.

描述（由申请人提供）：莱姆病是北半球最常见的媒介传播疾病，通过携带螺旋体伯氏疏螺旋体的蜱传播给人类。在蜱虫进食过程中营养物质突然涌入的情况下，细菌迅速重塑其表面，从而有效地建立哺乳动物感染。伯氏疏螺旋体宿主-病原体界面由脂蛋白主导，脂蛋白通过脂质修饰的半胱氨酸锚定在外膜的外小叶上。最近对小鼠感染模型的研究表明，外表面脂蛋白 C (OspC) 的表达对于早期感染的建立至关重要。本研究的总体目标是确定这一重要的螺旋体毒力因子的结构功能关系。 OspC 在细胞表面形成同型二聚体，其三维结构可分为三个不同的结构域：(i) 无序的 N 端“系链”肽，(ii) α 螺旋束“支架”，以及(iii)螺旋连接环的膜远端“圆顶”。然而，目前尚不清楚这三个结构域如何促进脂蛋白在细菌定植和传播中的生物学功能。 OspC 与回归热疏螺旋体 Vsp 蛋白属于同一蛋白折叠家族，参与免疫逃避和组织向性。事实上，Borrelia turicatae Vsp1 的晶体结构与 OspC 的偏差仅轻微，但最显着的是螺旋连接环。因此，我们假设 OspC 的圆顶残基对于 OspC 在早期感染期间的特定功能至关重要。此外，我们假设系链肽除了提供蛋白质分类信息（参见我们发表的工作）之外，还进化为通过优化其长度将毒力因子正确定位在细菌表面蛋白质组内。我们将在以下两个具体目标中测试这些假设： 1. 通过测试 Vsp 蛋白或 Vsp 支架-OspC dome 嵌合体是否可以在体外结合宿主分子中功能性地替代 OspC，确定 OspC dome 残基在定植和传播中的作用以及小鼠感染模型中的情况。最终，将确定功能所需的最小 OspC 圆顶结构域。 2. 通过测试现有的和新生成的 OspC 系链长度突变体的体外功能以及竞争性小鼠感染测定中的功能，确定 OspC 脂蛋白系链长度在毒力因子可及性和功能中的作用。这些研究将（i）在我们对螺旋体表面脂蛋白结构功能的研究中实现进一步的里程碑，（ii）显着增加我们对哺乳动物感染伯氏疏螺旋体期间早期事件的理解，以及（iii）产生可能最终转化为莱姆病的新型干预策略的设计。