Group II chaperonins are essential mediators of cellular protein folding in eukaryotes and archaea. These oligomeric protein machines, ∼1 megadalton, consist of two back-to-back rings encompassing a central cavity that accommodates polypeptide substrates1,2,3. Chaperonin-mediated protein folding is critically dependent on the closure of a built-in lid4,5, which is triggered by ATP hydrolysis6. The structural rearrangements and molecular events leading to lid closure are still unknown. Here we report four single particle cryo-electron microscopy (cryo-EM) structures of Mm-cpn, an archaeal group II chaperonin5,7, in the nucleotide-free (open) and nucleotide-induced (closed) states. The 4.3 Å resolution of the closed conformation allowed building of the first ever atomic model directly from the single particle cryo-EM density map, in which we were able to visualize the nucleotide and more than 70% of the side chains. The model of the open conformation was obtained by using the deformable elastic network modelling with the 8 Å resolution open-state cryo-EM density restraints. Together, the open and closed structures show how local conformational changes triggered by ATP hydrolysis lead to an alteration of intersubunit contacts within and across the rings, ultimately causing a rocking motion that closes the ring. Our analyses show that there is an intricate and unforeseen set of interactions controlling allosteric communication and inter-ring signalling, driving the conformational cycle of group II chaperonins. Beyond this, we anticipate that our methodology of combining single particle cryo-EM and computational modelling will become a powerful tool in the determination of atomic details involved in the dynamic processes of macromolecular machines in solution.
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Protein Data Bank
The three-dimensional cryo-EM density maps have been deposited into the EBI-MSD EMD database with accession codes: EMD-5137 (wild-type closed), EMD-5138 (lidless closed), EMD-5139 (wild-type open) and EMD-5140 (lidless open). The atomic models have been deposited in the Protein Data Bank as 3IYE (wild-type closed) and 3IYF (lidless open).
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We acknowledge the support of grants from the National Institutes of Health through the Nanomedicine Development Center Roadmap Initiative, Biomedical Technology Research Center for Structural Biology in National Center for Research Resources, Nanobiology Training Fellowship administered by the Keck Center of the Gulf Coast Consortia and the National Science Foundation.
Author Contributions J.Z. and J.J. collected the cryo-EM image data. J.Z. performed the image processing and reconstructions with the assistance of C.J.F. M.L.B. did the modelling and analysis of the closed state. G.F.S. did the model fitting for the open state. N.R.D. and S.R. designed the lidless mutation, the biochemical conditions for the cryo-EM experiments and performed all biochemical characterizations and experiments. S.J.L. advised on data processing and map filtering. J.Z., N.R.D., M.L.B., G.F.S., J.F. and W.C. interpreted the structural results. M.D., W.C. and J.Z. prepared all the movies. All authors contributed to the preparation of the manuscript.
This file contains Supplementary Methods, Supplementary References and Supplementary Figures 1-6 with Legends. (PDF 615 kb)
This movie shows 4.3 Å-resolution wild-type Mm-cpn cryo-EM density map in the closed state. (MP4 10638 kb)
This movie shows backbone model for the wild-type Mm-cpn closed-state subunit. (MP4 4018 kb)
This movie shows examples of visible sidechain densities in the wild-type closed state Mm-cpn cryo-EM density map. (MP4 10867 kb)
This movie shows different degrees of flexibility in the wild-type and lidless Mm-cpn in open and closed conformations. (MOV 1311 kb)
This movie shows simple geometric linear morphing between open and closed-state models of Mm-cpn to illustrate the spatial relationship of the two experimentally determined structures. This movie is not meant to represent the actual motion trajectory. (ZIP 19645 kb)
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Zhang, J., Baker, M., Schröder, G. et al. Mechanism of folding chamber closure in a group II chaperonin. Nature 463, 379–383 (2010). https://doi.org/10.1038/nature08701
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