Resolving the intoxication mechanism of botulinum neurotoxins using single molecule structural biology

利用单分子结构生物学解析肉毒杆菌神经毒素的中毒机制

• 批准号: 10717466
• 负责人: Mark E Bowen
• 金额: $ 42.74万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-20 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10717466
• 关键词: Biophysics; Botulinum Toxins; C-terminal; Cellular Membrane; Cholera Toxin; Classification; Clinical; Clinical Trials; Clostridium botulinum; Cosmetics; Crystallization; Data; Development; Diphtheria Toxin; Drug Kinetics; Electron Microscopy; Electrophysiology (science); Endosomes; Enzymes; Evolution; Family; Fluorescence; Fluorescence Resonance Energy Transfer; Goals; Heterogeneity; Imaging technology; Individual; Intoxication; Investigation; Ion Channel; Knowledge; Left; Life; Light; Link; Liposomes; Marketing; Membrane; Membrane Lipids; Methods; Modeling; Molecular; Molecular Conformation; N-terminal; Neurons; Neurotoxins; Peptide Hydrolases; Pharmacologic Substance; Physiological; Process; Protein Dynamics; Protein Engineering; Protein Isoforms; Proteins; Publishing; Reaction; Reagent; Serotyping; Structure; System; Techniques; Tetanus Toxin; Time; Toxin; Visit; biophysical techniques; cell type; clinically relevant; innovation; interest; man; member; molecular imaging; nanobodies; next generation; novel; novel strategies; preservation; prevent; receptor; receptor binding; sensor; simulation; single molecule; single-molecule FRET; stoichiometry; structural biology

Resolving the intoxication mechanism of botulinum neurotoxins using single molecule structural biology. The toxins produced by Clostridium botulinum are some of the deadliest known yet are also revered for their pharmaceutical utility. C. botulinum is classified into seven serotypes (A-G) based on the neurotoxins that they produce. Currently, pharmaceutical development has relied on botulinum neurotoxin type A1 (BoNT/A). However, botulinum neurotoxin type E (BoNT/E) is currently in clinical trials because it provides different pharmacokinetics, faster onset and shorter duration, which enable new treatment regimes. The BoNT proteins are members of the two-component, “AB toxin” family (e.g. tetanus, cholera, and diphtheria toxins), which inject a toxic cargo enzyme (part A) using a proteinaceous transmembrane delivery system (part B). As such, their structure and activity has been well studied. However, several fundamental open questions remain regarding the BoNT delivery mechanism, such as the number of toxins required to deliver the cargo. Additionally, while numerous structures have been solved of the dormant toxins, there is little structural information on the active delivery state(s). AB toxins deliver their cargo across cellular membranes, typically triggered by low pH, which causes structural changes of both parts A and B along with insertion into the membranes. The presence of aggregation at high protein concentrations and membranes provide many experimental challenges for techniques that rely on ensemble averaging. In contrast, single molecule fluorescence can observe individual proteins on single liposomes to revist these classic problems in AB toxin structural biology. These novel approaches will answer long-standing questions in the field and lead to new understanding of the differences between two clinically relevant isoforms.

使用单分子结构来解决肉毒神经毒素的近毒性机制 生物学。 梭状芽孢杆菌产生的毒素是最致命的毒素，但也被尊敬 他们的药品实用程序。基于 它们产生的神经毒素。目前，药物开发已在肉毒杆菌上获救 神经毒素A1型（BONT/A）。但是，肉毒杆菌神经毒素E型（BONT/E）目前处于临床状态 试验是因为它提供了不同的药代动力学，更快的发作和较短的持续时间，这使得 新的治疗方案。 BONT蛋白是两个组成部分“ AB Toxin”家族的成员（例如 破伤风，霍乱和白喉毒素），使用一种有毒的货物（A部分）使用 蛋白质跨膜输送系统（B部分）。因此，它们的结构和活动已经 井研究。但是，关于BONT交付仍然存在一些基本的开放问题 机制，例如运送货物所需的毒素数量。另外，虽然很多 结构已解决了休眠毒素的解决，活动的结构信息很少 交货状态。 AB毒素跨过细胞膜，通常由低pH触发， 这会导致两部分A和B的结构变化以及插入膜。 在高蛋白质浓度和膜上的聚集提供了许多实验 依靠合奏平均的技术面临的挑战。相反，单分子荧光可以 观察单个脂质体上的单个蛋白质，以抵抗AB毒素结构中的这些经典问题 生物学。这些新颖的方法将在现场回答长期存在的问题，并导致新的问题 了解两种临床相关同工型之间的差异。