ERASynBIO:ECF Express - An Orthogonal, Organism-independent Expression Platform Based on Extractoplasmic Function (ECF) Sigma Factors

ERASynBIO:ECF Express - 基于提取质功能 (ECF) Sigma 因子的正交、独立于生物体的表达平台

• 批准号: 1538946
• 负责人: Carol Gross
• 金额: $ 75万
• 依托单位: University of California-San Francisco
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-15 至 2018-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1538946&HistoricalAwards=false
• 关键词: ERASynBIO; ECF; Express; Orthogonal; Organism

The defining characteristics of synthetic biology are both the application of engineering principles to advance biological understanding and to develop new functions for useful purposes. One of the major challenges of synthetic biology is to modify or create new biological systems that are more efficient and reliable than natural organisms. This research project addresses one such challenge through the design and implementation of new regulatory modules that will control and optimize gene expression without wasting cellular resources. Significant broader impacts of this research are educating a next-generation workforce to support a growing bioeconomy, providing a new fundamental understanding of an important biological process, and generating new enabling tools for the wider synthetic biology community. This research takes advantage of a new process of using extracytoplasmic function sigma factors for synthetic biology applications. These extracytoplasmic function sigma factors (ECFs), when redesigned, can serve as robust gene regulatory switches capable of controlling diverse transcriptional activities. These factors have two conserved protein domains that recognize and bind to specific promoter sequences. They then recruit RNA-polymerase to initiate transcription from natural or synthetic promoter sites. The compositional simplicity of these factors are used for creating a series of synthetic sigma factor/promoter pairs tailored for specific regulatory needs. In this project, the investigators will create chimeric ECFs from natural ECFs to generate a large library of universal switches that are orthogonal in four model bacteria (E. coli, S. meliloti, B. subtilis and S. venezuelae). This will be accomplished by applying design strategies based on combinatorial synthesis, structure-guided mutational approaches and in vivo and in silico testing in complex genetic circuits. This results will provide new tools to enable biological design via synthetic biology. This project is the US collaborative component of a project funded through the ERASynBIO EU-US transnational funding mechanism.

合成生物学的定义特征是应用工程原理来促进生物学理解并开发用于有用目的的新功能。合成生物学的主要挑战之一是修改或创建比自然生物体更高效、更可靠的新生物系统。该研究项目通过设计和实施新的调控模块来解决这样的挑战，这些模块将控制和优化基因表达而不浪费细胞资源。这项研究的显着更广泛的影响是教育下一代劳动力以支持不断增长的生物经济，提供对重要生物过程的新的基本理解，并为更广泛的合成生物学界提供新的支持工具。这项研究利用了将胞质外功能西格玛因子用于合成生物学应用的新过程。这些胞质外功能西格玛因子（ECF）经过重新设计后，可以作为强大的基因调控开关，能够控制不同的转录活动。这些因子具有两个保守的蛋白质结构域，可识别并结合特定的启动子序列。然后，他们招募 RNA 聚合酶从天然或合成的启动子位点启动转录。这些因子的组成简单，用于创建一系列针对特定监管需求定制的合成西格玛因子/启动子对。 在该项目中，研究人员将从天然 ECF 中创建嵌合 ECF，以生成在四种模型细菌（大肠杆菌、苜蓿芽孢杆菌、枯草芽孢杆菌和委内瑞拉链球菌）中正交的大型通用开关库。这将通过应用基于组合合成、结构引导突变方法以及复杂遗传电路中的体内和计算机测试的设计策略来实现。这一结果将为通过合成生物学进行生物设计提供新的工具。该项目是通过 ERASynBIO 欧盟-美国跨国融资机制资助的项目的美国合作部分。