UKRI/BBSRC-NSF/BIO: Developing large serine integrases as tools for constructing and manipulating synthetic replicons

UKRI/BBSRC-NSF/BIO：开发大型丝氨酸整合酶作为构建和操作合成复制子的工具

• 批准号: 2223480
• 负责人: Phoebe Rice
• 金额: $ 158.33万
• 依托单位: University of Chicago
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-12-01 至 2026-11-30
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2223480&HistoricalAwards=false
• 关键词: UKRI; BBSRC; NSF; BIO; Developing

This collaborative work develops an expanded genetic toolkit to facilitate asking fundamental questions about how assemblies of genes work together to perform natural or synthetic functions. This toolkit is intended to be flexible and useful for the broad variety of creative genetic manipulations envisioned (past and future) by biologists, including the modular construction and convenient editing of programmable genetic circuits. The core of this toolkit is a family of enzymes termed “Large serine integrases (LSIs)” that mediate precise DNA rearrangements. These rearrangements can be reversed by the addition of a second protein called a recombination directionality factor (RDF). However, RDFs have been difficult to identify in natural sources, and how they work is not well understood. This work uses new approaches to address both of those problems. The PIs are developing ways to introduce high school students to the excitement of seeing and playing with the 3D structures of DNA, working with community groups to discuss the potential power of modern genetic tools and to encourage enrolment to university STEM-related subjects especially among women and underrepresented groups, and interfacing with the biotechnology industry to improve curriculum for industry-bound students.This collaborative study between US investigator Rice from the University of Chicago and UK investigator Olorunniji from Liverpool John Moores University brings together two groups with long-standing expertise in serine recombinases. The Olorunniji group’s expertise in synthetic biology and molecular biology is complemented by the Rice group’s expertise in structural biology. Their combined efforts will significantly expand and innovate the LSI-RDF toolkit that is available for use in synthetic cell studies. The project will: (1) identify novel integrase - RDF pairs with useful properties using both new and established computational approaches, followed by characterization in in vivo functional assays; (2) use the tools of structural biology to understand how RDFs interacts with their cognate LSIs, providing a solid foundation for protein engineering; (3) develop novel methods to alleviate the current bottleneck in identifying RDFs and (4) add new functionality to the toolbox by engineering RDFs to be light-responsive for rapid-response reaction reversal without the need for chemical inducers.This collaborative US/UK project is supported by the US National Science Foundation (NSF) and the UK Biotechnology and Biological Sciences Research Council (BBSRC), where NSF funds the US investigator and BBSRC funds the partners in the UK.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.

这项合作工作开发了一个扩展的遗传工具包，以促进询问有关基因组装如何协同工作以执行自然或合成功能的基本问题。该工具包旨在为生物学家设想的各种创造性遗传操作（过去和未来）提供灵活和有用的，包括模块化结构和可编程遗传电路的方便编辑。这个工具包的核心是一个被称为“大丝氨酸整合酶（LSIs）”的酶家族，它介导精确的DNA重排。这些重排可以通过添加第二种称为重组方向性因子（RDF）的蛋白质来逆转。然而，rdf很难在自然资源中识别，而且它们的工作原理也没有得到很好的理解。这项工作使用了新的方法来解决这两个问题。pi正在开发各种方法，向高中生介绍看到和玩DNA 3D结构的兴奋，与社区团体合作，讨论现代遗传工具的潜在力量，鼓励大学注册stem相关科目，特别是在女性和代表性不足的群体中，并与生物技术行业对接，以改进面向行业学生的课程。来自芝加哥大学的美国研究者Rice和来自利物浦约翰摩尔斯大学的英国研究者Olorunniji进行了这项合作研究，汇集了两个在丝氨酸重组酶方面具有长期专业知识的小组。Olorunniji小组在合成生物学和分子生物学方面的专业知识与Rice小组在结构生物学方面的专业知识相辅相成。他们的共同努力将极大地扩展和创新可用于合成细胞研究的LSI-RDF工具包。该项目将：(1)使用新的和已建立的计算方法识别具有有用特性的新型整合酶- RDF对，然后进行体内功能分析表征；(2)利用结构生物学的工具，了解RDFs如何与其同源lsi相互作用，为蛋白质工程提供坚实的基础；(3)开发新的方法来缓解目前识别rdf的瓶颈；(4)通过设计rdf，使其具有光响应性，从而在不需要化学诱导剂的情况下进行快速反应逆转，从而为工具箱增加新的功能。这个英美合作项目由美国国家科学基金会（NSF）和英国生物技术和生物科学研究委员会（BBSRC）支持，其中NSF资助美国研究者，BBSRC资助英国的合作伙伴。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。