NIH Research Festival
The Hsp70 chaperone system in eukaryotes is involved in maintaining protein homeostasis and protects the cell from various types of stress. Hsp70 prevents protein misfolding and promotes proper folding of nascent polypeptides and proteins. The homologue of Hsp70 in E. coli is DnaK. DnaK chaperone activity is dependent on two co-chaperones: a J-domain protein to assist in ATP hydrolysis and a nucleotide exchange factor (NEF) to exchange ADP with ATP. In E. coli, there are two cytosolic J-domain proteins DnaJ and CbpA that collaborate with DnaK and the E. coli NEF, GrpE. DnaJ and CbpA complement each other in the other’s absence in vivo, indicating overlapping activities and functions. Both DnaJ and CbpA contain a highly homologous J-domain involved in interaction and collaboration with DnaK. Additionally, DnaJ contains a cysteine-rich region involved in coordination of two zinc ions; this region is absent in CbpA. To better understand how DnaJ and CbpA differ in their co-chaperone activities, we tested them for the ability to reactivate heat-aggregated GFP in collaboration with DnaK and GrpE. We found that under the same reaction conditions, DnaJ reactivated GFP at a faster rate than CbpA. We also tested DnaJ and CbpA for the ability to interact with DnaK in a protein-protein interaction assay. DnaJ appeared to bind DnaK more tightly than CbpA. These results suggest a functional difference between DnaJ and CbpA in collaboration and interaction with DnaK.
Scientific Focus Area: Molecular Biology and Biochemistry
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