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Collaborative Research: Engineering MS2 Capsid Assembly Using Systematic Fitness Landscapes

合作研究：使用系统适应度景观设计 MS2 衣壳组装

基本信息

批准号：
2043973
负责人：
Danielle Tullman Ercek
金额：
$ 50.04万
依托单位：
Northwestern University
依托单位国家：
美国
项目类别：
Standard Grant
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=2043973&HistoricalAwards=false
关键词：
Collaborative Research Engineering MS2 Capsid

项目摘要

Much of the biological world is made up building blocks, known as proteins, which can assemble into complex structures at scales not visible to the naked eye. The proposed work explores the self-assembly of a structure made of proteins called a virus-like particle. Understanding how the building blocks of viruses come together could be important for developing therapeutics and vaccines to fight them. The objective of this work is to uncover the rules for how this virus particle assembles and how it can be modified to improve its function without disrupting its structure. Outreach efforts will be pursued that include engagement and training of undergraduate and graduate students, and engagement with K-12 students and the general public via demonstrations in schools and at science festivals, discussions at local libraries and museums, and creation and release of an episode about this work on a top-ranked podcast series for children ages 6-12.Virus-like particles are an excellent model system to study protein self-assembly. These closed-shell protein containers tend to be composed of a single protein and can be expressed to high titers in bacteria. They also have the useful property of encapsulating their own genetic information. This proposal takes advantage of this property to study both the advantageous and deleterious effects of mutations on the self-assembly properties of virus-like particles using an efficient new directed evolution technique. Importantly, this technique uses a single step to generate libraries of the particles in which all sequence positions are systematically substituted with every natural amino acid. These libraries can then be exposed to various selection pressures, enabling identification of particles with desirable physical or biochemical properties via the encapsulated RNA in survivors of the selection. Using a variety of mutagenesis and functional selections, this work aims to achieve two specific objectives. First, it will elucidate the critical requirements for assembly of the virus-like particle derived from bacteriophage MS2. Second, it will improve the chemical reactivity and stability of the particle so it can remain intact following modifications for any desired application.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

生物世界的大部分是由被称为蛋白质的积木组成的，这些蛋白质可以按肉眼看不到的规模组装成复杂的结构。这项拟议的工作探索了一种由蛋白质组成的结构的自组装，这种结构被称为病毒样颗粒。了解病毒的组成成分是如何结合在一起的，对于开发对抗它们的疗法和疫苗可能很重要。这项工作的目的是揭示这种病毒颗粒如何组装的规则，以及如何对其进行修改以在不破坏其结构的情况下改善其功能。将继续开展外联工作，包括本科生和研究生的参与和培训，以及通过在学校和科学节上的演示、在当地图书馆和博物馆的讨论以及为6-12岁的儿童制作和发布一集关于这项工作的顶级播客系列，与K-12学生和公众接触，病毒样颗粒是研究蛋白质自我组装的极佳模型系统。这些闭壳蛋白质容器往往由单一蛋白质组成，并可在细菌中高滴度表达。它们还具有封装自己的遗传信息的有用特性。这项建议利用这一性质，利用一种有效的新的定向进化技术来研究突变对病毒样颗粒自组装特性的有利和有害影响。重要的是，这项技术使用一个步骤来生成粒子库，其中所有序列位置都被每种天然氨基酸系统地取代。然后，这些文库可以暴露在各种选择压力下，从而能够通过选择幸存者体内的封装RNA来鉴定具有所需物理或生化特性的颗粒。利用各种诱变和功能选择，这项工作旨在实现两个特定的目标。首先，它将阐明组装来自噬菌体MS2的病毒样颗粒的关键要求。其次，它将提高颗粒的化学反应性和稳定性，使其在针对任何期望的应用进行修改后可以保持完好。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。