Multi-state recognition pathway of the intrinsically disordered protein kinase inhibitor by protein kinase A

  1. Cristina Olivieri
  2. Yingjie Wang
  3. Geoffrey C Li
  4. Manu Veliparambil Subrahmanian
  5. Jonggul Kim
  6. Benjamin R Stultz
  7. Matthew Neibergall
  8. Fernando Porcelli
  9. Joseph M Muretta
  10. David D T. Thomas
  11. Jiali Gao
  12. Donald K Blumenthal
  13. Susan S Taylor
  14. Gianluigi Veglia  Is a corresponding author
  1. University of Minnesota, United States
  2. Shenzhen Bay Laboratory, China
  3. Vanderbilt University, United States
  4. UT Southwestern Medical Center, United States
  5. Medical College of Wisconsin, United States
  6. Bethel University, United States
  7. University of Tuscia, Italy
  8. Beijing University Graduate School, China
  9. University of Utah, United States
  10. University of California, San Diego, United States

Abstract

In the nucleus, the spatiotemporal regulation of the catalytic subunit of cAMP-dependent protein kinase A (PKA-C) is orchestrated by an intrinsically disordered protein kinase inhibitor, PKI, which recruits the CRM1/RanGTP nuclear exporting complex. How the PKA-C/PKI complex assembles and recognizes CRM1/RanGTP is not well understood. Using NMR, SAXS, fluorescence, metadynamics, and Markov model analysis, we determined the multi-state recognition pathway for PKI. After a fast binding step in which PKA-C selects PKI's most competent conformations, PKI folds upon binding through a slow conformational rearrangement within the enzyme's binding pocket. The high-affinity and pseudo-substrate regions of PKI become more structured and the transient interactions with the kinase augment the helical content of the nuclear export sequence, which is then poised to recruit the CRM1/RanGTP complex for nuclear translocation. The multistate binding mechanism featured by PKA-C/PKI complex represents a paradigm on how disordered, ancillary proteins (or protein domains) are able to operate multiple functions such as inhibiting the kinase while recruiting other regulatory proteins for nuclear export.

Data availability

All NMR data will be deposited as NMR-Star File in the BMRB. Fluorescence data from Figures 5 and S5 will be deposited to the University of Minnesota repository site: https://conservancy.umn.edu/

The following data sets were generated

Article and author information

Author details

  1. Cristina Olivieri

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6957-6743
  2. Yingjie Wang

    Chemical Biology, Shenzhen Bay Laboratory, Shenzhen, China
    Competing interests
    The authors declare that no competing interests exist.
  3. Geoffrey C Li

    Biochemistry, Vanderbilt University, Nashville, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5035-5916
  4. Manu Veliparambil Subrahmanian

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Jonggul Kim

    Biophysics, UT Southwestern Medical Center, Dallas, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4624-7848
  6. Benjamin R Stultz

    Medicine, Medical College of Wisconsin, Milwaukee, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Matthew Neibergall

    Chemistry, Bethel University, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Fernando Porcelli

    DIBAF, University of Tuscia, Viterbo, Italy
    Competing interests
    The authors declare that no competing interests exist.
  9. Joseph M Muretta

    Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. David D T. Thomas

    Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8822-2040
  11. Jiali Gao

    School of Chemical Biology and Technology, Beijing University Graduate School, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  12. Donald K Blumenthal

    Pharmacology and Toxicology, University of Utah, Salt Lake City, United States
    Competing interests
    The authors declare that no competing interests exist.
  13. Susan S Taylor

    Department of Pharmacology, University of California, San Diego, La Jolla, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7702-6108
  14. Gianluigi Veglia

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    For correspondence
    vegli001@umn.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2795-6964

Funding

National Institute of General Medical Sciences (GM 100310)

  • Gianluigi Veglia

National Institute of General Medical Sciences (GM 46736)

  • Jiali Gao

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

Reviewing Editor

  1. Lewis E Kay, University of Toronto, Canada

Publication history

  1. Received: January 30, 2020
  2. Accepted: April 27, 2020
  3. Accepted Manuscript published: April 27, 2020 (version 1)
  4. Version of Record published: May 18, 2020 (version 2)
  5. Version of Record updated: May 26, 2020 (version 3)

Copyright

© 2020, Olivieri 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. Cristina Olivieri
  2. Yingjie Wang
  3. Geoffrey C Li
  4. Manu Veliparambil Subrahmanian
  5. Jonggul Kim
  6. Benjamin R Stultz
  7. Matthew Neibergall
  8. Fernando Porcelli
  9. Joseph M Muretta
  10. David D T. Thomas
  11. Jiali Gao
  12. Donald K Blumenthal
  13. Susan S Taylor
  14. Gianluigi Veglia
(2020)
Multi-state recognition pathway of the intrinsically disordered protein kinase inhibitor by protein kinase A
eLife 9:e55607.
https://doi.org/10.7554/eLife.55607

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