Novel Protein Folding Pathways for Protein Salvage and Recycling.

摘要

The goals for this project were as follows: 1. Demonstrate long term dispersion of aggregation-prone proteins with low temperature-active, mutant chaperones. 2. Develop refolding and salvage systems for maintaining proteins during long term incubation using chaperones with folding ability. Archaea are primitive microorganisms placed at the base of the Tree of Life, with greatly reduced genomes and proteomes compared to many mesophiles. In hyperthermophilic Archaea, the chaperonin (Cpn60) is the only ATP dependent protein folding complex is a 1 mDa molecular machine formed by two stacked rings of subunits. We have characterized the folding cycle in this machine and have created mutations that modulate its operating temperature to allow for efficient salvage of proteins. Using these chaperones we have accomplished the following: Manipulation of client protein range of chaperones, adjustment of E. coli's growth temperature, improved recombinant protein production, dismantling of amyloid proteins and inclusion bodies. Evidently, in vivo proteostasis enhancement systems resulted in enhanced high temperature growth and rapid folding of recombinant proteins. The discovery of reformulation of insoluble proteins opens new avenues for protein salvage.