Structural Basis for Autoinhibition of the EphB2 Receptor Tyrosine Kinase by the Unphosphorylated Juxtamembrane Region

Abstract

The Eph receptor tyrosine kinase family is regulated by autophosphorylation within the juxtamembrane region and the kinase activation segment. We have solved the X-ray crystal structure to 1.9 A resolution of an autoinhibited, unphosphorylated form of EphB2 comprised of the juxtamembrane region and the kinase domain. The structure, supported by mutagenesis data, reveals that the juxtamembrane segment adopts a helical conformation that distorts the small lobe of the kinase domain, and blocks the activation segment from attaining an activated conformation. Phosphorylation of conserved juxtamembrane tyrosines would relieve this autoinhibition by disturbing the association of the juxtamembrane segment with the kinase domain, while liberating phosphotyrosine sites for binding SH2 domains of target proteins. We propose that the autoinhibitory mechanism employed by EphB2 is a more general device through which receptor tyrosine kinases are controlled.

Cell–cell signaling via Eph receptors and ephrins

“…This brings about phosphorylation of the cytoplasmic Eph (as well as B-class ephrin) domains and downstream signaling initiation [3]. Kinase activation is controlled by the phosphorylation of residues in the activation loop and the juxtamembrane segment, which affect the inter-lobe(subdomain) dynamics of the kinase domain [31][32][33][34][35]. The minimal ligand-binding domain of the receptor is in blue and the ephrin receptor-binding domain is in pink.…”

“…In some RTKs, including the Eph, Kit, Flt, platelet-derived growth-factor beta (PDGFβ) and TrkB receptors, the juxtamembrane region is also involved in regulation of the kinase activity [31,32]. The crystal structure of the intracellular region of EphB2 [33] revealed that the unphosphorylated juxtamembrane region forms a well-ordered structure interacting with the N-terminal lobe of the kinase, presumably causing the distortion of a key α-helix and leading to kinase inactivation. Phosphorylation of the juxtamembrane tyrosine residues would push this region away via electrostatic repulsion, relieving the structural constraints that distort the active site.…”

FLT3: ITDoes matter in leukemia

“…Recently, structural and mutational analysis of EphB2, a member of the Eph receptor tyrosine kinase subfamily, has provided some insight into this issue. 94 In the inactive EphB2 receptor, components of the juxtamembrane domain, including two tyrosine residues (Y604 and Y610), are closely associated with an alpha helix of the N-terminal lobe of the kinase domain. This association prevents the activation loop within the kinase domain from assuming an active conformation.…”

Imatinib sensitivity as a consequence of a CSF1R-Y571D mutation and CSF1/CSF1R signaling abnormalities in the cell line GDM1

“…Interaction of Y571 with a conserved glutamic acid (E576) within the kinase domain is thought to contribute to stabilization of the receptor in an inactive conformation, 26 and its disruption by Y571 phosphorylation contributes to kinase activation. In more distantly related TKs such as the insulin receptor 27 and ephrin B2 28 receptors, the disruption of equivalently placed tyrosines and glutamic acid residues by tyrosine phosphorylation has also been implicated in kinase activation.…”