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: 10307559
• 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/10307559
• 关键词: Address; Anaerobic Bacteria; Assimilations; Bacteria; Biochemistry; Bioenergetics; Biogenesis; Bioinformatics; Biology; Catalysis; Complex; Conceptions; 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; base; cofactor; congenital infection; energy balance; 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.

项目概要/摘要 尽管梅毒作为人类瘟疫在历史上具有重要意义，但它仍然是最严重的疾病之一 了解所有人类感染。这是严格的研究限制的直接结果 历史上无法在体外连续培养梅毒螺旋体（Tp）。与更多的背离 与传统方法相比，大约 15 年前，我们开始了一项大胆的基于结构生物学的举措 表征 Tp 的脂蛋白 (LP)、对膜生物学、生物能学和至关重要的分子 Tp 的中间代谢，作为解开 Tp 机械进化“秘密”的一种手段 感染和梅毒发病机制。这种先进的研究途径已经成为非常成功的 发现平台，产生了许多高度新颖的发现，包括建立了许多新细菌 分子范式。例如，我们发现了一种新型双功能FAD焦磷酸酶/FMN Tp 中的转移酶；这反过来又使我们发现了 Tp 中的翻译后蛋白黄素酰化途径 周质，产生表面上影响细胞氧化还原反应的黄素蛋白。然后我们得到 Tp 编码非典型黄素基红细菌固氮 (RNF) 型氧化还原泵的证据， 可能代表了 Tp 膜电化学梯度、氧化还原之间长期缺失的联系 平衡、ATP 生成和产乙酸能量守恒途径。从历史上看，Tp 一直是 认为不对这样的系统进行编码。我们对基于黄素的氧化还原系统的争论不仅解决了 Tp 存在许多长期无法解释的代谢困境，但它也引发了范式转变 现在将 Tp 确定为黄素营养缺陷型。我们还证明了 TP0572，一个假定的 FMN- 依赖性铁还原酶，被 Ftp (TP0796) 黄素酰化，这可能是 Tp 的重要先决条件 还原性铁同化途径。此外，预测的胞质黄素蛋白必须发挥突出作用 保护 Tp 免受氧化应激并维持 NAD+/NADH 平衡。这些集体 有观点认为，在没有醌的情况下，ATP 产生的潜力有限，Tp 已经进化了 “以黄素为中心的代谢生活方式”，以满足人类感染的代谢需求。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（目标 1）、还原性铁同化和 Fe-S 蛋白生物发生（目标 2）以及氧化还原 平衡/能量守恒（通过产乙酸）（目标 3）。我们还将评估小分子 针对 Tp 黄素营养缺陷型的抑制剂作为潜在的新研究工具和/或新工具 针对 Tp 和其他致病性螺旋体的抗菌药物（目标 4）。综上所述，本项目应 阐明 Tp 如何进化以利用黄素作为其隐形基础的关键特征 致病性，可能导致阻止人类感染的新策略。