ME: Collaborative Research: Deployment of Enhanced Stress Responses to Improve Recombinant Expression Systems

该项目的目标是改进重组表达系统。主要目标是发现和表征超嗜热微生物中促进蛋白质折叠和亚基组装的途径。这些途径的组成部分将被插入重组宿主细胞中，以在重组表达过程中预先诱导应激反应，并增强重组蛋白的稳定性和溶解性。基于研究者先前研究的方法和结果，具体目标是：(1)确定超嗜热性细菌——炽热焦球菌（Pyrococcus furiosus）热休克调控中23个基因编码的相互作用蛋白的功能；(2)鉴定耐热性热休克蛋白的功能复合物，并在大肠杆菌、酵母和哺乳动物细胞中表达，增强这些细胞的耐久性；(3)确定伴侣蛋白组合对完整细胞和无细胞转录-翻译系统中蛋白高水平重组表达的影响。

The objective of this project is to improve recombinant expression systems. The major goals are to discover and characterize pathways that facilitate protein folding and subunit assembly in hyperthermophiles. The components of these pathways will be inserted into recombinant host cells to preempt induction of stress responses during recombinant expression, and to enhance the stabilization and solubilization of recombinant protein. Building on the approach and findings of the investigators previous research, the specific objectives are: (1) to determine the functions of interacting proteins encoded by the 23 genes in the heat shock regulon of the hyperthermophile, Pyrococcus furiosus, (2) to identify functional complexes of thermostable heat shock proteins and express them in E. coli, yeast and mammalian cells to enhance the durability of these cells, and (3) to determine the effects of combinations of chaperonins on high level recombinant expression of proteins in intact cells as well as cell-free transcription-translation systems.