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Functional and structural characterization of the kinase insert and the carboxy terminal domain in VEGF receptor 2 activation
Author(s) -
Manni Sandro,
Kisko Kaisa,
Schleier Thomas,
Missimer Jack,
BallmerHofer Kurt
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.14-256206
Subject(s) - protein kinase domain , microbiology and biotechnology , tyrosine kinase , kinase insert domain receptor , receptor tyrosine kinase , chemistry , kinase , tropomyosin receptor kinase c , platelet derived growth factor receptor , biology , signal transduction , biochemistry , receptor , vascular endothelial growth factor a , vascular endothelial growth factor , cancer research , vegf receptors , growth factor , gene , mutant
Vascular endothelial growth factors (VEGFs) regulate blood and lymphatic vessel development and homeostasis. VEGF receptor 2 (VEGFR‐2) is the major receptor involved in vasculogenesis and angiogenesis and regulates endothelial cell survival, migration, and mitogenesis. Ligand‐mediated receptor dimerization instigates transmembrane signaling, thereby promoting activation of the intracellular kinase domain. The intracellular part of the receptor comprises the juxtamembrane domain, the catalytic kinase domain, the kinase insert domain (KID), and the carboxy terminal domain (CD). Here we show that the CD inhibits VEGFR‐2 activity in the absence of ligand, whereas the KID, particularly a tyrosine residue in this domain (Y951), is indispensable for downstream signaling by the activated kinase. Because of the lack of crystallographic data for the complete kinase domain, we applied size‐exclusion chromatography, multiangle laser scattering, analytical ultracentrifugation, and small‐angle X‐ray scattering to build and functionally validate structural models. Our data show substantial conformational changes of the kinase when it is switched from the inactive, unphosphorylated state to the active, phosphorylated state. Finally, we structurally characterized recombinantly produced protein complexes between VEGFR‐2 and T cell‐specific adapter protein, a molecule involved in downstream signaling by VEGFR‐2.—Manni, S., Kisko, K., Schleier, T., Missimer, J., Ballmer‐Hofer, K., Functional and structural characterization of the kinase insert and the carboxy terminal domain in VEGF receptor 2 activation. FASEB J. 28, 4914–4923 (2014). www.fasebj.org