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Article

  • The EMBO Journal (2006) 25, 4686 - 4696
  • doi:10.1038/sj.emboj.7601315

Published online: 14 September 2006

A change in conformational dynamics underlies the activation of Eph receptor tyrosine kinases

Silke Wiesner1,a, Leanne E Wybenga-Groot2,a, Neil Warner2,3, Hong Lin1, Tony Pawson2,3, Julie D Forman-Kay1,4 and Frank Sicheri2,3

  1. Structural Biology and Biochemistry, Hospital for Sick Children, Toronto, Ontario, Canada
  2. Program in Systems Biology, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada
  3. Department of Molecular and Medical Genetics, University of Toronto, Toronto, Ontario, Canada
  4. Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada

Correspondence to:

Frank Sicheri, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Ave., Toronto, Ontario, Canada M5G 1X5. Tel.: +1 416 586 8471; Fax: +1 416 586 8869; E-mail: sicheri@mshri.on.ca

Julie D Forman-Kay, Hospital for Sick Children, 555 University Ave., Toronto, Ontario, Canada M5G 1X8. Tel.: +1 416 813 5358; Fax: +1 416 813 5022; E-mail: forman@sickkids.ca

Tony Pawson, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Ave., Toronto, Ontario, Canada M5G 1X5. Tel.: +1 416 586 8262; Fax: +1 416 586 8869; E-mail: pawson@mshri.on.ca

aThese authors contributed equally to this work

Received 21 April 2006; Accepted 8 August 2006

Eph receptor tyrosine kinases (RTKs) mediate numerous developmental processes. Their activity is regulated by auto-phosphorylation on two tyrosines within the juxtamembrane segment (JMS) immediately N-terminal to the kinase domain (KD). Here, we probe the molecular details of Eph kinase activation through mutational analysis, X-ray crystallography and NMR spectroscopy on auto-inhibited and active EphB2 and EphA4 fragments. We show that a Tyr750Ala gain-of-function mutation in the KD and JMS phosphorylation independently induce disorder of the JMS and its dissociation from the KD. Our X-ray analyses demonstrate that this occurs without major conformational changes to the KD and with only partial ordering of the KD activation segment. However, conformational exchange for helix alphaC in the N-terminal KD lobe and for the activation segment, coupled with increased inter-lobe dynamics, is observed upon kinase activation in our NMR analyses. Overall, our results suggest that a change in inter-lobe dynamics and the sampling of catalytically competent conformations for helix alphaC and the activation segment rather than a transition to a static active conformation underlies Eph RTK activation.

  • Keywords:

    • activation mechanism,
    • crystal structure,
    • Eph receptor tyrosine kinase,
    • NMR spectroscopy