Structure and Function of Treponema pallidum Lipoproteins

梅毒螺旋体脂蛋白的结构和功能

• 批准号: 8626308
• 负责人: MICHAEL V. NORGARD
• 金额: $ 57.66万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2019-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8626308
• 关键词: ATP-Binding Cassette Transporters; Bacteria; Biochemical; Biochemical Genetics; Biological; Biology; Cell physiology; Complex; Conceptions; Coupling; Data; Development; Flavin Mononucleotide; Flavins; Flavoproteins; Funding; Gene Cluster; Homeostasis; Homologous Gene; Immune Targeting; In Vitro; Intervention; Investigation; Ion Pumps; Life Style; Ligand Binding; Ligands; Lipoproteins; Membrane; Membrane Biology; Mission; Modality; Molecular; Nutritional; Operon; Order Spirochaetales; Organism; Oxidation-Reduction; Pathogenesis; Pathway interactions; Play; Potential Energy; Process; Property; Proteins; Research; Riboflavin; Role; Sexually Transmitted Diseases; Structure; Syphilis; System; Testing; Treponema pallidum; United States; Virulence Factors; Visit; Width; Work; antimicrobial drug; biophysical techniques; cofactor; design; genetic manipulation; member; novel; pathogen; periplasm; public health relevance; pyrophosphatase; receptor; structural biology; three dimensional structure; uptake

Project Summary/Abstract Treponema pallidum (Tp), the causative agent of syphilis, is one of most poorly understood of all bacterial pathogens. That Tp cannot be cultivated continuously in vitro has severely hampered progress towards understanding many of the complexities associated with Tp membrane biology and syphilis pathogenesis. Tp encodes a large number of membrane lipoproteins (LPs) that have widespread importance. Over the past funding interval, we have pursued a "structural biology" approach for characterizing the Tp LPs by solving their three-dimensional structures and then testing hypotheses about their possible functions. We now have compelling evidence that this unconventional approach represents a successful discovery platform. First, we have described in Tp a periplasmic transport system comprised of two LPs (TP0956/TP0957) that make up a TPR-protein-associated TRAP transport system. This system has many molecular features never before described for a bacterial transport apparatus. TP0956/TP0957 forms a complex that spans precisely the width of the Tp periplasm, and which contains a hydrophobic pore; we shall attempt to determine the ligand carried by this transport complex (Aim 3). We also have identified an essential ABC-type of riboflavin transporter (RfuABCD) in pathogenic spirochetes. This type of riboflavin transport system has never been previously described in bacteria, but it has prompted a re-examination of the potential importance of flavin-containing cofactors and flavoproteins to Tp biology. We also have discovered in Tp a novel FAD pyrophosphatase (TP0796/Ftp), the first such recognition of this type of enzymatic activity in any bacterium; the discovery has further implications for flavin utilization/homeostasis in Tp and, along with our other findings, engenders a new conception that Tp has evolved a "flavin-centric" lifestyle. Indeed, we have found that Tp encodes a homolog of RnfD (TP0151) that, in other bacteria, requires FMN as a co-factor and typically is a member of an RnfABCDGE operon for redox-driven ion pumps. Tp has been thought NOT to encode such as system, but we also have found a homolog of RnfC (TP0152) (within the gene cluster tp0147-tp0153). These combined findings engender strong supposition that Tp encodes a complete Rnf redox system, something that challenges existing dogma. Establishing that one or more of these Rnf-like proteins interact with each other will provide further evidence of an active Rnf-like redox system in Tp (Aim 2). A better understanding of this "flavin-centric" lifestyle for Tp may result in valuable new intervention strategies that target either riboflavin uptake or flavin utilization pathways, and could also provide further nutritional information leading to the eventual in vitro cultivation of Tp (Aim 1). Finally, all of our discoveries underscore the power of our "structure-to-function" approach, and thus we anticipate many more new advancements from discerning the structures of the Tp LPs (Aim 4.).

项目总结/摘要 梅毒螺旋体（Tp）是梅毒的病原体，是最不为人所知的疾病之一 细菌病原体TP不能在体外连续培养，这严重阻碍了进展 对了解许多复杂性与Tp膜生物学和梅毒 发病机制Tp编码大量的膜脂蛋白（LP），其具有广泛的生物学活性。 重要性在过去的资助期间，我们采用了“结构生物学”方法， 通过求解Tp LP的三维结构来表征Tp LP，然后检验假设 它们可能的功能。我们现在有令人信服的证据证明这种非传统的方法 这是一个成功的发现平台。首先，我们在Tp中描述了周质运输 由两个LP（TP 0956/TP 0957）组成的系统，构成TPR蛋白相关TRAP转运 系统该系统具有许多以前从未描述过的细菌转运的分子特征 设备. TP 0956/TP 0957形成一个精确跨越TP周质宽度的复合体，并且 它含有一个疏水孔;我们将试图确定这种运输所携带的配体 复合物（目标3）。我们还发现了一种必需的ABC型核黄素转运蛋白（RfuABCD）， 致病螺旋体这种类型的核黄素转运系统以前从未在文献中描述过。 但它促使人们重新审视含黄素辅因子的潜在重要性 和黄素蛋白到Tp生物学。我们还在Tp中发现了一种新的FAD焦磷酸酶 （TP 0796/Ftp），这是在任何细菌中首次识别这种类型的酶活性; 对Tp中的黄素利用/体内平衡有进一步的影响，沿着我们的其他发现， 产生了一个新的概念，即TP已经进化出一种“以黄素为中心”的生活方式。我确已发现， Tp编码RnfD（TP 0151）的同源物，其在其他细菌中需要FMN作为辅因子， 通常是用于氧化还原驱动离子泵的RnfABCDGE操纵子的成员。TP被认为不是 但是我们也发现了RnfC（TP 0152）的同源物（在基因簇内 tp0147-tp0153）。这些综合研究结果产生了强烈的假设，TP编码一个完整的Rnf 氧化还原系统，挑战现有的教条。建立一个或多个这些RNF样 蛋白质之间的相互作用将进一步证明TP中存在活性Rnf样氧化还原系统 (Aim 2）。更好地了解这种“黄素为中心”的生活方式为TP可能会导致有价值的新的 针对核黄素摄取或黄素利用途径的干预策略，还可以 提供进一步的营养信息，导致最终在体外培养的Tp（目标1）。最后所有 我们的发现强调了我们的“结构到功能”方法的力量，因此我们预计 从辨别Tp LP的结构中获得了更多的新进展（目标4.）。