Modified bacterial system for vivo direction evolution

用于体内定向进化的改良细菌系统

• 批准号: 6791132
• 负责人: Paul K. Olson
• 金额: $ 25.28万
• 依托单位: POTENTIA PHARMACEUTICALS, INC
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2006-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6791132
• 关键词: DNA replication; Escherichia coli; Neurospora; RNA directed DNA polymerase; bacteria; bacterial DNA; bacterial genetics; bacterial proteins; biochemical evolution; biotechnology; biotherapeutic agent; directed evolution; enzyme mechanism; evolution; fungal genetics; gene environment interaction; high throughput technology; site directed mutagenesis; technology /technique development

DESCRIPTION (provided by applicant): The goal of this proposal is to develop a new genetic system that will mimic the natural process of evolution for a selected subset of genes inside of a bacterial cell. Instead of an evolutionary time scale of years, evolution will proceed at an accelerated rate and data will be collected in a matter of days. In addition to providing a powerful system to study evolution in the laboratory, such a system will form the core of a directed evolution technology. Directed evolution is an approach that enables scientists to "tune" the properties of enzymes for specific applications. Its benefit has become so apparent that today most new enzymes undergo some type of directed evolution before being commercialized. However, all current implementations of directed evolution are conducted step-wise, with each step in the evolution process necessitating a substantial amount of time and manpower. A single cycle of evolution usually takes a few days and involves demanding technical work at each step. Because enzymes usually need to undergo many cycles of evolution, each application can take a substantial amount of time. This proposal will yield a continuous in vivo directed evolution platform requiring only minimal external input. Such an evolution platform will enable scientists to conduct more than 20 cycles of evolution in the course of a single day. It thus has the potential to be 10X faster than current evolution technologies, 10X cheaper, easier to implement, and yield proteins that will theoretically be better optimized than those produced today. Such a system will enable the optimization of proteins for various industrial applications (catalysts for pharmaceutical chemical synthesis is one example), but more importantly, it will be adapted to mammalian systems to enable the in vivo evolution of peptide-based therapeutics (such as antibodies, cyclic peptides, and enzymes). These therapeutics will address critical areas such as antiviral and antimicrobial drugs.

描述（由申请人提供）：该提案的目标是开发一种新的遗传系统，该系统将模拟细菌细胞内选定的基因子集的自然进化过程。进化的时间尺度不再是数年，而是加速进行，数据将在几天内收集完毕。除了提供一个强大的系统来在实验室研究进化之外，这样的系统还将构成定向进化技术的核心。定向进化是一种使科学家能够“调整”酶的特性以适应特定应用的方法。它的好处已经变得如此明显，以至于今天大多数新酶在商业化之前都会经历某种类型的定向进化。然而，目前所有定向进化的实现都是逐步进行的，进化过程中的每一步都需要大量的时间和人力。一个进化周期通常需要几天的时间，并且每一步都涉及艰巨的技术工作。由于酶通常需要经历多次进化循环，因此每次应用都可能需要大量时间。 该提案将产生一个仅需要最少外部输入的连续体内定向进化平台。这样的进化平台将使科学家能够在一天内进行20多个循环的进化。因此，它有可能比当前的进化技术快 10 倍、便宜 10 倍、更容易实施，并且产生的蛋白质理论上比现在生产的蛋白质更优化。这样的系统将能够优化各种工业应用的蛋白质（药物化学合成的催化剂就是一个例子），但更重要的是，它将适应哺乳动物系统，以实现基于肽的治疗剂（例如抗体）的体内进化、环肽和酶）。这些疗法将解决抗病毒和抗菌药物等关键领域。