The flavin-centric metabolic lifestyle of Treponema pallidum

梅毒螺旋体以黄素为中心的代谢生活方式

• 批准号: 10531598
• 负责人: MICHAEL V. NORGARD
• 金额: $ 54.39万
• 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 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10531598
• 关键词: Address; Bacteria; Biochemistry; Bioenergetics; Biogenesis; Bioinformatics; Biology; Catalysis; Complex; Conceptions; Cytoplasm; Electrochemistry; Electron Transport; Environment; Equilibrium; Flavin Mononucleotide; Flavins; Flavodoxin; Flavoproteins; Generations; Genomics; Homeostasis; Human; In Vitro; Infection; Intervention; Iron; Life Style; Link; Lipoproteins; Membrane; Membrane Biology; Metabolic; Metabolism; Modernization; Molecular; Molecular Biology; NADH; Nitrogen Fixation; Order Spirochaetales; Oxidation-Reduction; Oxidative Stress; Pathogenesis; Pathogenicity; Pathway interactions; Plague; Play; Process; Proteins; Pump; Quinones; Reaction; Research; Research Personnel; Rhodobacter; Riboflavin; Role; Sexually Transmitted Diseases; Syphilis; System; Tissues; Transferase; Treponema pallidum; United States; antimicrobial; auxotrophy; cofactor; congenital infection; enzyme pathway; genetic manipulation; iron (III) reductase; metalloenzyme; novel; periplasm; pyrophosphatase; small molecule inhibitor; structural biology; tool; uptake

Project Summary/Abstract Despite its historical importance as a plague on humankind, syphilis remains among the most poorly understood of all human infections. This is a direct result of severe research constraints imposed by the historic inability to cultivate Treponema pallidum (Tp) continuously in vitro. In a departure from more conventional approaches, about 15 years ago we embarked on a bold structural biology-based initiative to characterize Tp’s lipoproteins (LPs), molecules critical to the membrane biology, bioenergetics, and intermediary metabolism of Tp, as a means of unlocking the mechanistic evolutionary “secrets” of Tp infection and syphilis pathogenesis. This progressive research avenue has become a very successful discovery platform, yielding many highly novel findings, including establishing a number of new bacterial molecular paradigms. For example, we discovered a novel bi-functional FAD pyrophosphatase/FMN transferase in Tp; this, in turn, led us to identify a post-translational protein flavinylation pathway in Tp’s periplasm, yielding flavoproteins that ostensibly influence cellular redox reactions. We then obtained evidence for Tp encoding an atypical flavin-based Rhodobacter Nitrogen Fixation (RNF)-type redox pump, likely representing the longstanding missing link between Tp’s membrane electrochemical gradient, redox balance, ATP generation, and an acetogenic energy conservation pathway. Historically, Tp has been thought not to encode such systems. Our contention of a flavin-based redox system not only addresses a number of longstanding unexplained metabolic dilemmas for Tp, but it also engenders a paradigm shift by now establishing Tp as a flavin auxotroph. We also have demonstrated that TP0572, a putative FMN- dependent ferric reductase, is flavinylated by Ftp (TP0796), likely an essential prerequisite for Tp’s reductive iron assimilation pathway(s). In addition, predicted cytosolic flavoproteins must play prominently in protecting Tp from oxidative stress and in maintaining the balance of NAD+/NADH. These collective notions support that, with limited potential for ATP generation in the absence of quinones, Tp has evolved a “flavin-centric metabolic lifestyle” to fulfill its metabolic requirements for human infection. This project shall address three core metabolic features relevant to Tp’s flavin biology: protein flavinylation and flavoprotein biogenesis (Aim 1), reductive iron assimilation and Fe-S protein biogenesis (Aim 2), and redox balance/energy conservation (via acetogenesis) (Aim 3). We also shall evaluate a small-molecule inhibitor(s) targeting Tp’s flavin auxotrophy as a potential new research tool(s) and/or new antimicrobial(s) against Tp and other pathogenic spirochetes (Aim 4). Taken together, this project shall elucidate key features concerning how Tp has evolved to exploit flavins as an underpinning of its stealth pathogenicity, potentially leading to new strategies to thwart human infection.

项目摘要/摘要 尽管梅毒是对人类的瘟疫的历史重要性，但仍然是最糟糕的 了解所有人类感染。这是严重研究限制的直接结果 历史性无法在体外连续培养毛毛虫（TP）。偏离更多 常规方法，大约15年前，我们着手采取一项大胆的基于结构生物学的计划 特征TP的脂蛋白（LPS），对膜生物学，生物能学至关重要的分子 TP的中介代谢，作为解锁TP机械进化“秘密”的一种手段 感染和梅毒发病机理。这座进步的研究大道已成为非常成功的 发现平台产生了许多新颖的发现，包括建立许多新细菌 分子范式。例如，我们发现了一种新型的双功率焦磷酸酶/FMN TP中的转移酶；反过来，这导致我们确定了TP的翻译后蛋白黄素化途径 周期质，产生表面上会影响细胞氧化还原反应的黄素蛋白。然后我们获得了 TP编码基于非典型黄素的杜鹃杆菌（RNF）型氧化还原泵的证据， 也许代表了TP的膜电化学梯度之间的长期缺失的联系，氧化还原 平衡，ATP的产生和乙酸能源保护途径。从历史上看，TP一直是 认为不编码此类系统。我们对基于黄素的氧化还原系统的争论不仅解决了 TP的长期无法解释的代谢难题数量，但它也导致了范式的转变 现在将TP建立为黄素合子。我们还证明了TP0572，一种推定的FMN- 依赖的铁还原，通过FTP（TP0796）进行黄素化，这可能是TP的重要先决条件 还原铁同化途径。此外，预测的胞质黄素蛋白必须显着发挥 在保护TP免受氧化应激和维持NAD+/NADH的平衡方面。这些集体 注释支持的是，在没有奎因酮的情况下ATP产生的潜力有限，TP已经进化 一种“以黄素为中心的代谢生活方式”，以满足其对人类感染的代谢要求。这个项目 应解决与TP黄素生物学相关的三个核心代谢特征：蛋白质黄素化和 黄素蛋白生物发生（AIM 1），铁同化和Fe-S蛋白生物发生（AIM 2）和氧化还原 平衡/能量保存（通过乙酰生成）（AIM 3）。我们还将评估小分子 抑制剂（S）靶向TP的黄素合物作为潜在的新研究工具和/或新的 针对TP和其他致病性螺旋体的抗菌剂（AIM 4）。总之，这个项目应 阐明有关TP如何演变成其隐形的基础的关键功能 致病性，有可能导致阻止人类感染的新策略。