Ribosome structure determination from Apicomplexan parasites

顶复门寄生虫的核糖体结构测定

基本信息

批准号：
10726704
负责人：
Melissa Leger-Abraham
金额：
$ 25.43万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-05-24 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10726704
关键词：
Address Affinity Affinity Chromatography Animal Diseases Animals Anti-Bacterial Agents Antibiotics Antiparasitic Agents Apicomplexa Area Babesia Babesiosis Binding Biology Clinical Complex Cryoelectron Microscopy Cytoplasm Cytoplasmic Structures Cytosol Development Disease Drug resistance Elements Erythrocytes Event Fever Foundations Gene Expression Gene Expression Regulation Geographic Distribution Geography Goals Growth Hemagglutinin Human Immune system Immunocompromised Host Individual Infection Inorganic Phosphate Transporter Laboratories Life Life Cycle Stages Malaria Membrane Methods Mitochondria Molecular Molecular Biology Molecular Structure NMR Spectroscopy Names Organelles Organism Parasites Pathway interactions Pharmaceutical Preparations Plasmodium Plasmodium falciparum Preparation Property Protein Biochemistry Protein Biosynthesis Protocols documentation Public Health Research Proposals Resolution Ribosomal Proteins Ribosomal RNA Ribosomes Structure Symptoms Therapeutic Intervention Thick Tick-Borne Diseases Toxoplasma Toxoplasmosis Translations Vaccines Work X-Ray Crystallography antibiotic design climate change design experience experimental study forest human disease improved novel novel therapeutics particle posttranscriptional rational design structural biology suburb targeted agent targeted treatment tick-borne

项目摘要

PROJECT SUMMARY/ABSTRACT The Apicomplexa phylum contains many parasites that infect and cause disease in humans and animals, including the agents that cause malaria, toxoplasmosis, and babesiosis. Babesia is a tick-borne intracellular parasite that causes severe to fatal diseases, with broadening geographic distribution because of climate change and increasing suburban developments in forested areas. There is no vaccine against human babesiosis, and the current therapies have limited efficacy. Considering the complex life cycle of Babesia species, their survival depends on the precise control of their gene expression, which is mostly regulated post-transcriptionally (e.g., during protein synthesis). Babesia species have a unique, non-photosynthetic organelle named apicoplast, derived from a secondary endosymbiotic event. In addition to having ribosomes in its cytoplasm and mitochondria, the apicoplast contains its own ribosomes. Little is known about the structure of these ribosomes, although they are thought to be highly simplified, with prokaryotic-like components. They are inhibited by antibacterial drugs that have some limited antiparasitic activity. The absence of a molecular structure of any apicoplast ribosomes limits the rational design of antibiotics with improved antiparasitic properties. In the first aim of this research proposal, we will establish a Babesia divergens strain expressing an hemagglutinin (HA)- tagged triose phosphate transporter on the outer membrane of the apicoplast. This strategy will allow the affinity purification of a high-quality organelle preparation, followed by the purification of apicoplast ribosomes. In the second aim, we will define the components of apicoplast ribosomes and determine their structure using cryo- electron microscopy (cryo-EM). Our goal is thus to elucidate the fundamental molecular machinery that apicomplexan parasites use to synthesize proteins in the highly specialized apicoplast organelle. Our work will lay the foundation for studies that interrogate these parasites' gene regulation mechanisms and help inspire the design of better antiparasitic agents.

项目概要/摘要顶复门含有许多感染人类和动物并引起疾病的寄生虫，包括引起疟疾、弓形体病和巴贝斯虫病的病原体。巴贝斯虫是一种蜱传细胞内寄生虫导致严重甚至致命疾病的寄生虫，由于气候变化其地理分布范围不断扩大并增加森林地区的郊区开发。没有针对人类巴贝斯虫病的疫苗，并且目前的疗法疗效有限。考虑到巴贝虫物种复杂的生命周期，它们的生存取决于对其基因表达的精确控制，这主要是在转录后调控的（例如，蛋白质合成过程中）。巴贝虫属物种有一种独特的非光合作用细胞器，名为 apicoplast，源自次级内共生事件。除了细胞质中含有核糖体外，线粒体，顶质体含有自己的核糖体。人们对这些核糖体的结构知之甚少，尽管它们被认为是高度简化的，具有类似原核生物的成分。它们被抑制具有一定有限抗寄生虫活性的抗菌药物。不存在任何分子结构顶质体核糖体限制了具有改进的抗寄生虫特性的抗生素的合理设计。在第一个本研究计划的目的是，我们将建立表达血凝素 (HA) 的巴贝斯虫菌株 - 顶端质体外膜上标记的磷酸丙糖转运蛋白。该策略将允许亲和力纯化高质量的细胞器制剂，然后纯化顶质体核糖体。在第二个目标，我们将定义顶端质体核糖体的成分，并使用冷冻技术确定其结构电子显微镜（冷冻电镜）。因此，我们的目标是阐明基本的分子机制顶端复门寄生虫用于在高度特化的顶端质体细胞器中合成蛋白质。我们的工作将为探究这些寄生虫的基因调控机制的研究奠定基础，并有助于启发设计更好的抗寄生虫药物。