Structural transitions modulate the chaperone activities of Grp94

Author:

Amankwah Yaa S.12ORCID,Fleifil Yasmeen1,Unruh Erin13,Collins Preston1,Wang Yi2,Vitou Katherine1,Bates Alison1ORCID,Obaseki Ikponwmosa1ORCID,Sugoor Meghana1,Alao John Paul1,McCarrick Robert M.1,Gewirth Daniel T.4,Sahu Indra D.5,Li Zihai2ORCID,Lorigan Gary. A.13,Kravats Andrea N.13ORCID

Affiliation:

1. Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056

2. Pelotonia Institute for Immuno-Oncology, Division of Medical Oncology, The Ohio State University Comprehensive Cancer Center–Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, The Ohio State University, Columbus, OH 43210

3. Cell, Molecular, and Structural Biology Graduate Program, Miami University, Oxford, OH 45056

4. Hauptman-Woodward Medical Research Institute, Buffalo, NY 14203

5. Natural Sciences Division, Campbellsville University, Campbellsville, KY 42718

Abstract

Hsp90s are ATP-dependent chaperones that collaborate with co-chaperones and Hsp70s to remodel client proteins. Grp94 is the ER Hsp90 homolog essential for folding multiple secretory and membrane proteins. Grp94 interacts with the ER Hsp70, BiP, although the collaboration of the ER chaperones in protein remodeling is not well understood. Grp94 undergoes large-scale conformational changes that are coupled to chaperone activity. Within Grp94, a region called the pre-N domain suppresses ATP hydrolysis and conformational transitions to the active chaperone conformation. In this work, we combined in vivo and in vitro functional assays and structural studies to characterize the chaperone mechanism of Grp94. We show that Grp94 directly collaborates with the BiP chaperone system to fold clients. Grp94’s pre-N domain is not necessary for Grp94–client interactions. The folding of some Grp94 clients does not require direct interactions between Grp94 and BiP in vivo, suggesting that the canonical collaboration may not be a general chaperone mechanism for Grp94. The BiP co-chaperone DnaJB11 promotes the interaction between Grp94 and BiP, relieving the pre-N domain suppression of Grp94’s ATP hydrolysis activity. In structural studies, we find that ATP binding by Grp94 alters the ATP lid conformation, while BiP binding stabilizes a partially closed Grp94 intermediate. Together, BiP and ATP push Grp94 into the active closed conformation for client folding. We also find that nucleotide binding reduces Grp94’s affinity for clients, which is important for productive client folding. Alteration of client affinity by nucleotide binding may be a conserved chaperone mechanism for a subset of ER chaperones.

Funder

HHS | NIH | National Institute of General Medical Sciences

National Science Foundation

Publisher

Proceedings of the National Academy of Sciences

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