Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.12 (PKG)
 2,515 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The major receptor protein for cyclic GMP (cGMP) in smooth muscle is the cGMP-dependent protein kinase (cGMP kinase). The more abundant I alpha isoform (subunit M(r) congruent to 78,000) of this enzyme mediates the effects of cGMP to relax contracted vascular smooth muscle preparations. In this study, we have addressed the hypothesis that the cGMP kinase is anchored to intracellular proteins which might serve to target cGMP kinase to protein substrates. Using a gel overlay technique, immunoprecipitation, and a fluorescence binding assay for cGMP kinase, we have identified vimentin as a high-affinity and specific binding protein for cGMP kinase. Binding of cGMP kinase to vimentin is reversible and stoichiometric (one cGMP kinase dimer/vimentin dimer) with a KD of approximately 49 nM. The site of high-affinity binding between cGMP kinase and vimentin did not appear to be localized to the catalytic domain of the kinase since vimentin phosphorylated by cGMP kinase and peptide substrates for cGMP kinase did not compete for high-affinity binding. Neither the proteolytically-derived 69-kDa catalytic fragment nor the 8-kDa N-terminal fragment bound vimentin with high affinity, suggesting that the cGMP kinase dimer was necessary for the interaction. Vimentin was readily phosphorylated in vitro with the dimer, but not the monomeric 69-kDa catalytic fragment even though the monomeric 69-kDa fragment was catalytically active toward other substrates such as histone F2b and peptides. This suggests that the high-affinity interaction between cGMP kinase and vimentin occurs at the N-terminal region, thus allowing the interaction between the phosphorylation site of vimentin and the catalytic site of cGMP kinase to occur.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:High-affinity binding and localization of the cyclic GMP-dependent protein kinase with the intermediate filament protein vimentin. 802 8

The cytoskeleton plays a central role for the integration of biochemical and biomechanical signals across the cell required for complex cellular functions. Recent studies indicate that the intermediate filament vimentin is necessary for endothelial cell morphogenesis e.g. in the context of leukocyte transmigration. Here, we present evidence, that the scaffold provided by vimentin is essential for VASP localization and PKG mediated VASP phosphorylation and thus controls endothelial cell migration and proliferation. Vimentin suppression using siRNA technique significantly decreased migration velocity by 50% (videomicroscopy), diminished transmigration activity by 42.5% (Boyden chamber) and reduced proliferation by 43% (BrdU-incorporation). In confocal microscopy Vimentin colocalized with VASP and PKG in endothelial cells. Vimentin suppression was accompanied with a translocation of VASP from focal contacts to the perinuclear region. VASP/Vimentin and PKG/Vimentin colocalization appeared to be essential for proper PKG mediated VASP phosphorylation because we detected a diminished expression of PKG and p(Ser239)-VASP in vimentin-suppressed cells, Furthermore, the induction of VASP phosphorylation in perfused arteries was markedly decreased in vimentin knockout mice compared to wildtypes. A link is proposed between vimentin, VASP phosphorylation and actin dynamics that delivers an explanation for the important role of vimentin in controlling endothelial cell morphogenesis.
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PMID:Vimentin expression influences flow dependent VASP phosphorylation and regulates cell migration and proliferation. 2038 23