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

本项目的目的是改进重组表达系统。 主要目标是发现和表征促进超嗜热菌中蛋白质折叠和亚基组装的途径。 将这些途径的组分插入重组宿主细胞中，以在重组表达期间抢先诱导应激反应，并增强重组蛋白的稳定性和溶解性。 本论文在前人研究的基础上，具体目标是：（1）确定热激调节子中23个基因编码的相互作用蛋白的功能;（2）鉴定热稳定热激蛋白的功能复合物，并在大肠杆菌中表达。大肠杆菌、酵母和哺乳动物细胞，以增强这些细胞的耐久性，和（3）确定伴侣蛋白的组合对完整细胞以及无细胞转录-翻译系统中蛋白质的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-freetranscription-translation systems.