A novel biotherapeutic expression platform in bacteria

细菌中的新型生物治疗表达平台

• 批准号: 6880357
• 负责人: MATTHEW P DELISA
• 金额: $ 10万
• 依托单位: VYBION, INC.
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-02-01 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6880357
• 关键词: Escherichia coli; acidity /alkalinity; arginine; bacterial proteins; bioreactors; biotechnology; biotherapeutic agent; drug design /synthesis /production; hydrogen transport; membrane transport proteins; molecular chaperones; protein engineering; protein folding; protein transport; recombinant proteins; secretory protein; temperature

DESCRIPTION (provided by applicant): The overall objective of the proposal is to develop a novel protein expression platform by capitalizing on the remarkable properties of the recently discovered twin-arginine translocation (Tat) pathway of bacteria. With over 300 therapeutic proteins currently in various stages of clinical trials, the road to a healthier future will require new methods for producing safer and less expensive recombinant proteins. For this purpose, the recently discovered twin-arginine translocation (Tat) pathway will be utilized as a novel recombinant protein expression platform in Escherichia coli. Towards this objective, Phase I encompasses the following specific aims: 1) to define the secretory capacity of the bacterial Tat transporter; 2) to optimize Tat secretion efficiency via chaperone co-expression strategies; and 3) to scale-up the Tat-based protein production process. This research is expected to not only result in the development a novel platform for bacterial protein expression but will also facilitate a deeper understanding of a poorly understood biological mechanism. Phase II will entail a much broader effort to study and improve Tat transport efficiency including, for instance, directed evolution strategies to engineer "hyperactive" Tat transporters, cell engineering methods to optimize the host for Tat expression and leader peptide optimization studies. In addition, we expect to fully develop scale-up methods for large-scale Tat expression by using optimized carbon-source feed strategies, as well as temperature, dissolved oxygen and pH optimization.

描述（由申请人提供）：该提案的总体目的是通过利用最近发现的细菌最近发现的双精氨酸易位（TAT）途径的显着特性来开发新型的蛋白质表达平台。目前有300多种治疗蛋白处于临床试验的各个阶段，通往更健康的未来的道路将需要新的方法来生产更安全，更便宜的重组蛋白。为此，最近发现的双精氨酸易位（TAT）途径将被用作大肠杆菌中新型的重组蛋白表达平台。朝向这个目标，第一阶段涵盖了以下特定目的：1）定义细菌TAT转运蛋白的分泌能力； 2）通过伴侣共表达策略来优化TAT分泌效率； 3）扩大基于TAT的蛋白质生产过程。预计这项研究不仅会导致开发细菌蛋白表达的新型平台，而且还将促进对知识熟悉的生物学机制的更深入了解。第二阶段将需要更广泛的努力来研究和提高TAT的运输效率，例如，包括针对“过度活跃” TAT转运蛋白的指示进化策略，用于优化宿主的TAT表达和领导者肽优化研究的细胞工程方法。此外，我们期望通过使用优化的碳源饲料策略以及温度，溶解氧和pH优化来完全开发大规模TAT表达的规模方法。