Highly Dynamic Interactions Maintain Kinetic Stability of the ClpXP Protease During the ATP-Fueled Mechanical Cycle
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Author(s)
Sello, Jason K.
Sauer, Robert T.
Amor, Alvaro Jorge
Baker, Tania
Schmitz, Karl R.
Date Issued
March 2016
Journal
ACS Chemical Biology
Publisher
American Chemical Society (ACS)
Citation
Amor, Alvaro J.; Schmitz, Karl R.; Sello, Jason K.; Baker, Tania A. and Sauer, Robert T. “Highly Dynamic Interactions Maintain Kinetic Stability of the ClpXP Protease During the ATP-Fueled Mechanical Cycle.” ACS Chemical Biology 11, no. 6 (June 17, 2016): 1552–1560. © 2016 American Chemical Society (ACS)
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Author's final manuscript
Abstract
The ClpXP protease assembles in a reaction in which an ATP-bound ring hexamer of ClpX binds to one or both heptameric rings of the ClpP peptidase. Contacts between ClpX IGF-loops and clefts on a ClpP ring stabilize the complex. How ClpXP stability is maintained during the ATP-hydrolysis cycle that powers mechanical unfolding and translocation of protein substrates is poorly understood. Here, we use a real-time kinetic assay to monitor the effects of nucleotides on the assembly and disassembly of ClpXP. When ATP is present, complexes containing single-chain ClpX assemble via an intermediate and remain intact until transferred into buffers containing ADP or no nucleotides. ATP binding to high-affinity subunits of the ClpX hexamer prevents rapid dissociation, but additional subunits must be occupied to promote assembly. Small-molecule acyldepsipeptides, which compete with the IGF loops of ClpX for ClpP-cleft binding, cause exceptionally rapid dissociation of otherwise stable ClpXP complexes, suggesting that the IGF-loop interactions with ClpP must be highly dynamic. Our results indicate that the ClpX hexamer spends almost no time in an ATP-free state during the ATPase cycle, allowing highly processive degradation of protein substrates.
MIT Department
Massachusetts Institute of Technology. Department of Biology
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DOI of Published Version
https://doi.org/10.1021/acschembio.6b00083
