PlzA, cyclic-di-GMP and the enzootic cycle for Lyme disease

PlzA、环二 GMP 和莱姆病的地方性循环

基本信息

批准号：
10608622
负责人：
Christopher Davies
金额：
$ 74.6万
依托单位：
VIRGINIA COMMONWEALTH UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-11-18 至 2027-10-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10608622
关键词：
Alleles Amino Acids Bacteria Binding Binding Proteins Binding Sites Biological Biological Assay Biological Process Biopsy Borrelia burgdorferi CISH gene Cell physiology Chemotaxis Co-Immunoprecipitations DNA Data Electrophoretic Mobility Shift Assay Environment Gene Expression Genes Genome Growth Hand In Vitro Incidence Infection Ixodes Ligand Binding Ligands Lyme Disease Mammals Mediating Midgut Molecular Molecular Chaperones Morphology Mus Mutate Mutation Mutation Analysis Nature Northern Blotting Organ Outcome Outcome Study Periodicity Plasmids Process Property Proteins Public Health RNA RNA Binding RNA annealing RNA immunoprecipitation sequencing Regulation Regulator Genes Sequence Analysis Series Site Site-Directed Mutagenesis Structure Structure-Activity Relationship Surface Surface Plasmon Resonance Tertiary Protein Structure Testing Tick-Borne Diseases Ticks Tissues Transgenic Organisms Translations Validation Work cell motility comparative diguanylate cyclase environmental change enzootic gene replacement lyme pathogenesis mutant overexpression posttranscriptional protein protein interaction seroconversion transmission process

项目摘要

Project summary: Lyme disease is the most common tick-borne disease in the northern hemisphere and its incidence is steadily increasing. The ability of the Lyme disease spirochetes to adapt to changing environmental conditions is dependent in part on regulation mediated by the secondary messenger molecule, cyclic-di-GMP (c-di-GMP). This proposal will identify the critical functional determinants of PlzA, the sole identified c-di-GMP binding protein produced by all Lyme disease spirochete isolates. Based on the recently determined atomic structure of PlzA which revealed that PlzA belongs to the unique xPilZ domain class of c-di-GMP binding proteins and data demonstrating that PlzA has RNA chaperone activities, we will perform a comprehensive analysis to dissect structure-function relationships of both apo and holo forms of the protein. Based on comparative sequence and structural analyses, surface-exposed amino acid residues will be targeted for site-directed mutagenesis. The impact of these mutations on PlzA structure, c-di-GMP binding, RNA winding, RNA unwinding, and protein-protein interactions will be assessed in vitro. Based on these analyses, a series of B. burgdorferi transgenic strains in which wild-type plzA is replaced with plzA genes that encode PlzA proteins with altered activity will be generated. The ability of each strain to infect and transmit between ticks and mammals will be determined. These analyses will define the functional domains and biological mechanisms by which apo and holo PlzA regulate cellular processes required for the completion of the enzootic cycle.

项目概要：莱姆病是北方地区最常见的蜱传疾病半球，其发病率正在稳步增加。莱姆病螺旋体的能力适应不断变化的环境条件部分依赖于监管第二信使分子，环二 GMP (c-di-GMP)。该提案将确定关键的 PlzA 的功能决定因素，PlzA 是所有莱姆病产生的唯一鉴定的 c-di-GMP 结合蛋白疾病螺旋体分离株。基于最近确定的 PlzA 原子结构揭示 PlzA 属于 c-di-GMP 结合蛋白的独特 xPilZ 结构域类别，并且数据表明 PlzA 具有 RNA 伴侣活性，我们将进行全面的分析来剖析蛋白质的 apo 和全息形式的结构-功能关系。基于比较序列和结构分析，表面暴露的氨基酸残基将成为定点诱变的目标。这些突变对 PlzA 结构的影响， c-di-GMP 结合、RNA 缠绕、RNA 解旋和蛋白质-蛋白质相互作用将体外评估。基于这些分析，一系列伯氏疏螺旋体转基因菌株，其中野生型 plzA 被编码活性改变的 PlzA 蛋白的 plzA 基因取代生成的。每种菌株在蜱虫和哺乳动物之间感染和传播的能力将是决定。这些分析将通过以下方式定义功能域和生物机制： apo 和 Holo PlzA 调节完成地方性动物病所需的细胞过程循环。