Tandem repeats of highly bioluminescent NanoLuc are refolded noncanonically by the Hsp70 machinery

In this study, we further investigate these properties through in vitro bulk experiments. Similar to monomeric Nluc, engineered Nluc dyads and triads proved to be highly biolumines cent. Using the bioluminescence signal as the proxy for their structural integrity, we determined that heat-denatured Nluc dyads and triads can be efficiently refolded by theE. coli Hsp70 chaperone system, which comprises DnaK, DnaJ, and GrpE. In contrast to previous studies with other substrates, we observed that Nluc repeats can be efficiently refolded by DnaK and DnaJ, even in the absence of GrpE co-chaperone. Taken together, our study offers a new powerful substrate for chaperone research and raises intriguing questions about the Hsp70 mechanisms, particularly in the context of structurally diverse proteins.

Source: Protein Science - January 23, 2024 Category: Biochemistry Authors: Dimitra Apostolidou, Pan Zhang, Devanshi Pandya, Kaden Bock, Qinglian Liu, Weitao Yang, Piotr E. Marszalek Tags: RESEARCH ARTICLE Source Type: research