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Crystal structures of the JAK2 pseudokinase domain and the pathogenic mutant V617F

Nature Structural & Molecular Biology volume 19, pages 754–759 (2012)Cite this article

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Abstract

The protein tyrosine kinase JAK2 mediates signaling through numerous cytokine receptors. JAK2 possesses a pseudokinase domain (JH2) and a tyrosine kinase domain (JH1). Through unknown mechanisms, JH2 regulates the catalytic activity of JH1, and hyperactivating mutations in the JH2 region of human JAK2 cause myeloproliferative neoplasms (MPNs). We showed previously that JAK2 JH2 is, in fact, catalytically active. Here we present crystal structures of human JAK2 JH2, including both wild type and the most prevalent MPN mutant, V617F. The structures reveal that JH2 adopts the fold of a prototypical protein kinase but binds Mg-ATP noncanonically. The structural and biochemical data indicate that the V617F mutation rigidifies α-helix C in the N lobe of JH2, facilitating trans-phosphorylation of JH1. The crystal structures of JH2 afford new opportunities for the design of novel JAK2 therapeutics targeting MPNs.

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Figure 1: Crystal structure of JAK2 JH2.
Figure 2: Cis- versus trans-autophosphorylation of JAK2 JH2.
Figure 3: Comparison of JH2 V617F and wild-type structures.
Figure 4: Basal activation state of JAK2 mutants in mammalian cells.

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Acknowledgements

This work was supported in part by US National Institute of Allergy and Infectious Diseases grant R21 AI095808 (S.R.H.), the Medical Research Council of Academy of Finland (O.S.), the Sigrid Juselius Foundation (O.S.), the Finnish Cancer Foundation (O.S.), the Competitive Research Funding and Centre of Laboratory Medicine of the Tampere University Hospital (O.S.), the Maire Lisko Foundation (O.S.) and the Tampere Tuberculosis Foundation (O.S.). US National Cancer Institute training grant T32 CA009161 supported R.M.B. Financial support for beam line X25 at the National Synchrotron Light Source, Brookhaven National Laboratory, comes principally from the US Department of Energy, National Center for Research Resources (P41 RR012408) and National Institute of General Medical Sciences (P41 GM103473). We thank E. Koskenalho for technical assistance, V. DiGiacomo for peptide modeling, A. Héroux for X-ray data collection, W.T. Miller, M. Mohammadi and K. Gnanasambandan for critical reading of the manuscript and M. Eck (Dana-Farber Cancer Institute, Boston, Massachusetts, USA) for discussions and for providing coordinates before publication.

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Author notes

  1. Rajintha M Bandaranayake and Daniela Ungureanu: These authors contributed equally to this work.

Authors and Affiliations

  1. Structural Biology Program, Kimmel Center for Biology and Medicine at the Skirball Institute, New York University School of Medicine, New York, New York, USA

    Rajintha M Bandaranayake & Stevan R Hubbard

  2. Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, New York, USA

    Rajintha M Bandaranayake & Stevan R Hubbard

  3. Institute of Biomedical Technology, University of Tampere and Tampere University Hospital, Tampere, Finland

    Daniela Ungureanu & Olli Silvennoinen

  4. D.E. Shaw Research, New York, New York, USA

    Yibing Shan & David E Shaw

  5. Center for Computational Biology and Bioinformatics, Columbia University, New York, New York, USA

    David E Shaw

Authors
  1. Rajintha M Bandaranayake
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  2. Daniela Ungureanu
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Contributions

R.M.B., crystallographic studies and manuscript preparation; D.U., in-cell biochemical studies and manuscript preparation; Y.S. and D.E.S., molecular dynamics simulations and manuscript preparation; O.S., project supervisor and manuscript preparation; S.R.H., project supervisor and principal manuscript author.

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Correspondence to Olli Silvennoinen or Stevan R Hubbard.

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Bandaranayake, R., Ungureanu, D., Shan, Y. et al. Crystal structures of the JAK2 pseudokinase domain and the pathogenic mutant V617F. Nat Struct Mol Biol 19, 754–759 (2012). https://doi.org/10.1038/nsmb.2348

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