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Analysis of Reconstituted Tripartite Complex Supports Avidity-based Recruitment of Hsp70 by Substrate Bound J-domain Protein
Abstract
Hsp70 are ubiquitous, versatile molecular chaperones that cyclically interact with substrate protein(s). The initial step requires synergistic interaction of a substrate and a J-domain protein (JDP) cochaperone, via its J-domain, with Hsp70 to stimulate hydrolysis of its bound ATP. This hydrolysis drives conformational changes in Hsp70 that stabilize substrate binding. However, because of the transient nature of substrate and JDP interactions, this key step is not well understood. Here we leverage a well characterized Hsp70 system specialized for iron-sulfur cluster biogenesis, which like many systems, has a JDP that binds substrate on its own. Utilizing an ATPase-deficient Hsp70 variant, we isolated a Hsp70-JDP-substrate tripartite complex. Complex formation and stability depended on residues previously identified as essential for bipartite interactions: JDP-substrate, Hsp70-substrate and J-domain-Hsp70. Computational docking based on the established J-domain-Hsp70(ATP) interaction placed the substrate close to its predicted position in the peptide-binding cleft, with the JDP having the same architecture as when in a bipartite complex with substrate. Together, our results indicate that the structurally rigid JDP-substrate complex recruits Hsp70(ATP) via precise positioning of J-domain and substrate at their respective interaction sites - resulting in functionally high affinity (i.e., avidity). The exceptionally high avidity observed for this specialized system may be unusual because of the rigid architecture of its JDP and the additional JDP-Hsp70 interaction site uncovered in this study. However, functionally important avidity driven by JDP-substrate interactions is likely sufficient to explain synergistic ATPase stimulation and efficient substrate trapping in many Hsp70 systems.
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Authors (14)
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Marcin Jelen
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Igor Grochowina
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Aneta Grabinska-Rogala
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Szymon Ciesielski
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Katarzyna Dabrowska
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Bartlomiej Tomiczek
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Wojciech Delewski
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Brenda Schilke
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Michal Dadlez
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Rafal Dutkiewicz
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Elizabeth Craig
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Jaroslaw Marszalek
Cite as
Full text
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- Accepted or Published Version
- DOI:
- Digital Object Identifier (open in new tab) 10.1016/j.jmb.2023.168283
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Details
- Category:
- Articles
- Type:
- artykuły w czasopismach
- Published in:
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JOURNAL OF MOLECULAR BIOLOGY
no. 435,
ISSN: 0022-2836 - Language:
- English
- Publication year:
- 2023
- Bibliographic description:
- Jelen M., Grochowina I., Grabinska-Rogala A., Ciesielski S., Dabrowska K., Tomiczek B., Nierzwicki Ł., Delewski W., Schilke B., Czub J., Dadlez M., Dutkiewicz R., Craig E., Marszalek J.: Analysis of Reconstituted Tripartite Complex Supports Avidity-based Recruitment of Hsp70 by Substrate Bound J-domain Protein// JOURNAL OF MOLECULAR BIOLOGY -Vol. 435,iss. 21 (2023), s.168283-
- DOI:
- Digital Object Identifier (open in new tab) 10.1016/j.jmb.2023.168283
- Sources of funding:
-
- COST_FREE
- Verified by:
- Gdańsk University of Technology
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