Conformational and oligomeric states of SPOP from small-angle X-ray scattering and molecular dynamics simulations

  1. F Emil Thomasen
  2. Matthew J Cuneo
  3. Tanja Mittag
  4. Kresten Lindorff-Larsen  Is a corresponding author
  1. University of Copenhagen, Denmark
  2. St. Jude Children's Research Hospital, United States

Abstract

Speckle-type POZ protein (SPOP) is a substrate adaptor in the ubiquitin proteasome system, and plays important roles in cell-cycle control, development, and cancer. SPOP forms linear higher-order oligomers following an isodesmic self-association model. Oligomerization is essential for SPOP's multivalent interactions with substrates, which facilitate phase separation and localization to biomolecular condensates. Structural characterization of SPOP in its oligomeric state and in solution is, however, challenging due to the inherent conformational and compositional heterogeneity of the oligomeric species. Here, we develop an approach to simultaneously and self-consistently characterize the conformational ensemble and the distribution of oligomeric states of SPOP by combining small-angle X-ray scattering (SAXS) and molecular dynamics simulations. We build initial conformational ensembles of SPOP oligomers using coarse-grained molecular dynamics simulations, and use a Bayesian/maximum entropy approach to refine the ensembles, along with the distribution of oligomeric states, against a concentration series of SAXS experiments. Our results suggest that SPOP oligomers behave as rigid, helical structures in solution, and that a flexible linker region allows SPOP's substrate binding domains to extend away from the core of the oligomers. Additionally, our results are in good agreement with previous characterization of the isodesmic self-association of SPOP. In the future, the approach presented here can be extended to other systems to simultaneously characterize structural heterogeneity and self-assembly.

Data availability

Code and data is available at https://github.com/KULL-Centre/_2022_Thomasen_SPOP. Simulation data is available at https://doi.org/10.17894/ucph.ef999f72-b5e8-45c4-835f-3e49619a0f91. Plasmids are available from Addgene (plasmid IDs 194115 and 194116).

The following data sets were generated

Article and author information

Author details

  1. F Emil Thomasen

    Department of Biology, University of Copenhagen, Copenhagem, Denmark
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2096-4873
  2. Matthew J Cuneo

    Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1475-6656
  3. Tanja Mittag

    Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, United States
    Competing interests
    Tanja Mittag, Tanja Mittag was a consultant for Faze Medicines, Inc..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1827-3811
  4. Kresten Lindorff-Larsen

    Department of Biology, University of Copenhagen, Copenhagen, Denmark
    For correspondence
    lindorff@bio.ku.dk
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4750-6039

Funding

Lundbeckfonden (R155-2015-2666)

  • Kresten Lindorff-Larsen

Novo Nordisk Fonden (NNF18OC0033950)

  • Kresten Lindorff-Larsen

National Institutes of Health (R01GM112846)

  • Tanja Mittag

American Lebanese Syrian Associated Charities

  • Tanja Mittag

Novo Nordisk Fonden (NNF18OC0032608)

  • Kresten Lindorff-Larsen

National Institutes of Health (P30GM133893)

  • Tanja Mittag

DOE Office of Biological and Environmental Research (KP1605010)

  • Tanja Mittag

National Institutes of Health (OD012331)

  • Tanja Mittag

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2023, Thomasen et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. F Emil Thomasen
  2. Matthew J Cuneo
  3. Tanja Mittag
  4. Kresten Lindorff-Larsen
(2023)
Conformational and oligomeric states of SPOP from small-angle X-ray scattering and molecular dynamics simulations
eLife 12:e84147.
https://doi.org/10.7554/eLife.84147

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https://doi.org/10.7554/eLife.84147

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