Toward Artificial Proteomes

走向人工蛋白质组

• 批准号: 1409402
• 负责人: Michael Hecht
• 金额: $ 98.57万
• 依托单位: Princeton University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2019-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1409402&HistoricalAwards=false
• 关键词: Toward; Artificial; Proteomes

A proteome can be defined as the entire collection of proteins in an organism. Thus, a proteome can be viewed as the complete set of molecular machines necessary to sustain a living organism. Because many different biochemical functions are required in any particular cell, the proteome of any single organism must include a wide range of proteins with diverse amino acid sequences, three-dimensional structures, and biochemical activities. Nonetheless, the full collection of all proteins in all proteomes that ever existed on earth constitutes a minuscule fraction of the sequences that are theoretically possible. Thus, despite billions of years of evolutionary sampling, the vast majority of sequence space remains unexplored. However, recent advances in synthetic biology, combinatorial methods, and protein design have made it possible to begin exploring sequences that have never been exposed to evolution. The proposed research aims to design and produce large collections of novel proteins that fold into well-defined 3D structures, and function in biologically relevant reactions. Completion of this work will represent a significant advance toward developing artificial proteomes that were not evolved by nature, but nonetheless support the growth of living organisms. The project will have broader significance in basic science and for applied technologies: Richard Feynman said, "What I cannot create, I do not understand." Thus, the creation of novel proteins will both test our knowledge, and enhance our understanding of protein biochemistry, biophysics, and molecular evolution. Design and construction of artificial proteomes will also impact applied science and biotechnology: Current biotechnology relies on protein sequences borrowed from nature, while future applications will benefit from de novo sequences that were not selected by nature, but are well-suited for industrial applications. This project provides excellent interdisciplinary research training opportunities. The investigator will present this work also to the public and discuss it in the classroom.Previous studies of proteins and proteomes were limited to sequences isolated (or modified) from natural systems. The proposed research will overcome this limitation by making available libraries of millions of novel proteins. Such collections will be 1000-fold larger than typical bacterial proteomes. In contrast to studies of natural proteomes, which reveal what was selected by nature, newly enabled studies of artificial proteomes will broaden our understanding beyond what evolved in nature, and will enable experiments that probe sequences, structures, and functions, which are not observed in natural biological systems, but nonetheless can occur in the realm of novel or synthetic biologies. The research will harness both combinatorial/ experimental and computational/theoretical approaches to pursue the following aims:-Design and construction of large collections of novel alpha-helical proteins.-Design and construction of large collections of novel beta-sheet proteins. Proteomes, whether natural or artificial, must contain both alpha and beta structures. -Development and implementation of a high throughput screen for folded structures. The quality of protein libraries will be enhanced by subjecting collections of computationally designed proteins to follow-up screens for structures that are soluble and stably folded.-Determination of 3-dimensional structures and stabilities of proteins from the novel proteome. Successful designs will produce sequences that fold into expected structures.-Isolation and evolution of novel proteins that are active in vitro and functional in vivo. Most importantly, collections of novel sequences will resemble proteomes if and only if they include proteins that are biochemically active and provide essential cellular functions.

蛋白质组可以定义为生物体中蛋白质的整个集合。 因此，可以将蛋白质组视为维持生物体所需的完整分子机器。 由于在任何特定细胞中都需要许多不同的生化功能，因此任何单个生物体的蛋白质组都必须包括各种具有多种氨基酸序列，三维结构和生化活性的蛋白质。尽管如此，在地球上存在的所有蛋白质组中所有蛋白质的完整收集构成了理论上可能的序列的微小分数。因此，尽管进行了数十亿年的进化采样，但绝大多数序列空间仍未开发。但是，合成生物学，组合方法和蛋白质设计的最新进展使得开始探索从未暴露于进化的序列。 拟议的研究旨在设计和生产大量新型蛋白质，这些蛋白质折叠成明确的3D结构，并在生物学相关的反应中起作用。 这项工作的完成将代表着发​​展自然界并非发展的人工蛋白质组织的重大进步，但仍支持生物的生长。该项目将在基础科学和应用技术中具有更大的意义：理查德·费曼（Richard Feynman）说：“我无法创建的东西，我不了解。” 因此，新型蛋白质的产生既可以测试我们的知识，又增强我们对蛋白质生物化学，生物物理学和分子进化的理解。 人工蛋白质组织的设计和构建还将影响应用科学和生物技术：当前的生物技术依赖于从自然界借来的蛋白质序列，而未来的应用将受益于自然界未选择的从头序列，但适合工业应用。该项目提供了出色的跨学科研究培训机会。 研究人员还将向公众介绍这项工作，并在课堂上进行讨论。对蛋白质和蛋白质组的预防研究仅限于从天然系统中分离（或修改）的序列。 拟议的研究将通过提供数百万个新型蛋白质的库来克服这一局限性。此类收集将比典型的细菌蛋白质组织大1000倍。 与揭示自然选择的天然蛋白质组织的研究相反，对人造蛋白质组的新实现研究将使我们的理解范围扩大到自然界发展的范围之外，并将实现探测序列序列，结构和功能的实验，这些实验在自然生物学系统中未观察到，但仍可以在新颖或合成生物的领域中发生。这项研究将利用组合/实验和计算/理论方法来追求以下目的： - 设计和构建大量新型α-螺旋蛋白。设计和建造大量新型Beta-sheet蛋白质。蛋白质组织，无论是天然还是人造，都必须包含α和β结构。 - 用于折叠结构的高吞吐量屏幕的开发和实现。通过将计算设计蛋白质的蛋白质的集合作为可溶性且稳定折叠的结构的后续筛选，将提高蛋白质文库的质量。确定了新型蛋白质组的三维结构和稳定性。 成功的设计将产生将序列折叠成期望结构的序列。在体外活性和实用的新型蛋白质的异化和演变。 最重要的是，新序列的集合将类似于蛋白质组，并且仅当它们包括生化活性并提供必要的细胞功能的蛋白质时。