Directed and adaptive evolution of photosynthetic systems

光合系统的定向和适应性进化

• 批准号: MR/Y011635/1
• 负责人: Tanai Cardona Londono
• 金额: $ 75.59万
• 依托单位: Queen Mary University of London
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FY011635%2F1
• 关键词: Directed; adaptive; evolution; photosynthetic; systems

Photosystems are the complex molecular assemblies of photosynthesis, they generate the energy that sustain nearly all the biosphere, directly powering the global fixation of about a hundred gigatons of carbon dioxide annually. Photosynthesis uses two photosystems working in series: photosystem II and photosystem I. Photosystem II harvests light to power the oxidation and decomposition of water into protons, electrons, and the oxygen we breathe. Photosystem I harvests light to generate the power needed to drive metabolic reactions, importantly, but not exclusively, carbon dioxide fixation. These properties make the photosystems amongst the most powerful enzymes in the history of life, both capable of driving their own difficult chemistry using light.My long-term vision is to use evolution-based methods to completely redesign the photosystems so that we can harness their properties to do useful chemistry beyond their naturally evolved function. I want to develop a technological platform that enables exquisite control of both photosystems to achieve bespoke multi-step light-driven oxidative or reductive biocatalysis. Eventually, I envision this platform linked to a research facility that allows for the rapid and high-throughput purification, characterisation, and production of these novel photosystems for a broad range of applications, from bioremediation to precision chemistry, all driven by light. Therefore, this technology can benefit and impact the biotechnology and chemical industry by creating new ways to perform clean chemical reactions.In this extension, I continue the work that was started in the first stage of the fellowship. My lab is currently developing, optimising, and characterising directed evolution approaches that target photosystem II to change its functional properties beyond its naturally evolved function. Directed evolution is an extremely versatile approach that is used to change the traits, or the activity of a given enzyme by exploiting evolution. It can be done simply by subjecting an organism through repeated cycles of selection under the conditions that favour the desired traits or by screening for the desired function, it can be enhanced by turbocharging mutational rates (hypermutation), it can be focused on a single gene of interest, parts of a gene, or multiple genes. Hypermutation can be done in vitro, where the genes are mutated in the test tube; or in vivo, where hypermutation occurs as the cells divide and replicate. My lab has a working in vitro system that we have already used to isolate several photosystem II variants harbouring a range of mutations and are about to deploy and in vivo CRISPR-based hypermutation system.The idea of applying directed evolution to engineer novel photosystems is without precedent. While directed evolution is a somewhat of a mature technology, it has not been extensively applied to complex enzymatic systems like the photosystems, if at all. In addition, the development of directed evolution approaches in photosynthetic organisms also lags compared with non-photosynthetic systems like E. coli and yeast. While an ambitious and challenging research endeavour, I've demonstrated that photosystem II remains evolvable and plastic in nature thanks to its structural modularity. Therefore, our approach harnesses this natural adaptability using directed evolution but accelerated to lab timescales.The overarching aim of this fellowship is to demonstrate that photosystem II is amenable to directed evolution, and to begin developing hypermutation and selection approaches to drive its evolution. The ultimate objective of this extension is to deliver proof-of-concept novel photosystems, or strains of cyanobacteria harbouring these novel photosystems, that could pave the wave towards scaling up and translating this vision into viable green biotechnologies.

光系统是光合作用的复杂分子组件，它们产生了几乎所有生物圈的能量，每年直接为全球固定大约一百吉型二氧化碳的全球固定。光合作用使用两个光系统串联的光系统：光系统II和光系统II。光系统II收获光，为水的氧化和分解为质子，电子和我们呼吸的氧气。光系统I收集光，以产生驱动代谢反应所需的功率，重要的是，但不是仅限于二氧化碳固定。这些特性使光系统在生命史上最强大的酶之一，既能够使用光线来驱动自己的困难化学反应。我的长期视觉是使用基于进化的方法来完全重新设计光系统，以便我们可以利用其特性来在其自然演化的功能之外进行有用的化学作用。我想开发一个技术平台，该平台能够对这两个光系统进行精美的控制，以实现定制的多步驱动氧化或还原性生物催化。最终，我设想了这个与研究设施相关的平台，该平台允许这些新型光系统的快速和高通量纯化，表征和生产，用于从生物修复到精密化学的广泛应用，全部由光驱动。因此，这项技术可以通过创建新的清洁化学反应方法来受益并影响生物技术和化学工业。在此扩展中，我继续从研究金的第一阶段开始的工作。我的实验室目前正在开发，优化和表征定向的进化方法，该方法以光化系统II为目标，以将其功能属性更改为自然发展的功能。定向进化是一种极其用途的方法，用于通过利用进化来改变特征或给定酶的活性。可以简单地通过在有利于所需性状的条件下的反复选择循环或筛选所需功能的条件下进行生物体来完成，它可以通过涡轮增压突变速率（超称）来增强，它可以集中于单个基因，基因的部分或多个基因的单个基因。可以在体外进行超突变，其中基因在试管中被突变。或体内，当细胞分裂和复制时，超突变发生。我的实验室具有一个工作的体外系统，我们已经用来隔离具有一系列突变的几种光系统II变体，即将部署和基于体内CRISPR的超伪装系统。将有方向的进化应用于工程师小说的光系统的想法是没有先例的。虽然导向的进化是一种成熟的技术，但它并未广泛应用于像光系统（如果有的话）等复杂的酶促系统。此外，与大肠杆菌和酵母这样的非光合合成系统相比，光合生物体中定向进化方法的发展也滞后。虽然这是一项雄心勃勃且充满挑战的研究努力，但我证明了光系统II由于其结构模块化而在本质上仍然可以发展和塑料。因此，我们的方法利用定向进化来利用这种自然的适应性，但加速到实验室时间表。该团契的总体目的是证明光系统II可以适合定向进化，并开始开发超名和选择方法来推动其进化。该扩展的最终目标是提供概念验证的新型光系统，或含有这些新型光系统的蓝细菌菌株，可以将波浪铺平到扩大并将这种视力转化为可行的绿色生物技术。