Rowland, Rhianna J., Heath, Richard, Maskell, Daniel et al. (6 more authors) (2023) Cryo-EM structure of SKP1-SKP2-CKS1 in complex with CDK2-cyclin A-p27KIP1. Scientific reports. 10718. ISSN 2045-2322
Abstract
p27KIP1 (cyclin-dependent kinase inhibitor 1B, p27) is a member of the CIP/KIP family of CDK (cyclin dependent kinase) regulators that inhibit cell cycle CDKs. p27 phosphorylation by CDK1/2, signals its recruitment to the SCFSKP2 (S-phase kinase associated protein 1 (SKP1)-cullin-SKP2) E3 ubiquitin ligase complex for proteasomal degradation. The nature of p27 binding to SKP2 and CKS1 was revealed by the SKP1-SKP2-CKS1-p27 phosphopeptide crystal structure. Subsequently, a model for the hexameric CDK2-cyclin A-CKS1-p27-SKP1-SKP2 complex was proposed by overlaying an independently determined CDK2-cyclin A-p27 structure. Here we describe the experimentally determined structure of the isolated CDK2-cyclin A-CKS1-p27-SKP1-SKP2 complex at 3.4 Å global resolution using cryogenic electron microscopy. This structure supports previous analysis in which p27 was found to be structurally dynamic, transitioning from disordered to nascent secondary structure on target binding. We employed 3D variability analysis to further explore the conformational space of the hexameric complex and uncovered a previously unidentified hinge motion centred on CKS1. This flexibility gives rise to open and closed conformations of the hexameric complex that we propose may contribute to p27 regulation by facilitating recognition with SCFSKP2. This 3D variability analysis further informed particle subtraction and local refinement approaches to enhance the local resolution of the complex.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | Funding Information: M.S. and R.F.T are current employees of Evotec SE and Thermo Fisher Scientific respectively. The other authors declare no competing interests. The authors declare no competing financial interest. Some work in the authors’ laboratory is supported by a research grant from Astex Pharmaceuticals. Funding Information: This research was supported by the Medical Research Council (Grant References MR/N009738/1 and MR/V029142/1). Cryogenic electron microscopy was carried out at the York Structural Biology Lab (Wellcome Trust grant number 206161/Z/17/Z) and Leeds Astbury Centre. At the University of Leeds, the FEI Titan Krios microscopes were funded by the University of Leeds (UoL ABSL award) and The Wellcome Trust (108466/Z/15/Z), with Falcon 4 and Selectris Falcon4 direct electron detector and microED packages (Q2 2021) upgrades funded by The Wellcome Trust (221524/Z/20/Z). We would like to thank S. Lea, D. Elmlund and H. Elmlund for their assistance during initial stages of this project. For the purpose of open access, the authors have applied a Creative Commons Attribution (CC BY) licence to any Author Accepted Manuscript version arising from this submission. Publisher Copyright: © 2023, The Author(s). |
Dates: |
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Institution: | The University of York |
Academic Units: | The University of York > Faculty of Sciences (York) > Chemistry (York) |
Funding Information: | Funder Grant number WELLCOME TRUST 206161/Z/17/Z |
Depositing User: | Pure (York) |
Date Deposited: | 18 Oct 2023 09:10 |
Last Modified: | 07 Jan 2025 00:17 |
Published Version: | https://doi.org/10.1038/s41598-023-37609-9 |
Status: | Published |
Refereed: | Yes |
Identification Number: | 10.1038/s41598-023-37609-9 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:204355 |
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Description: Cryo-EM structure of SKP1-SKP2-CKS1 in complex with CDK2-cyclin A-p27KIP1
Licence: CC-BY 2.5