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Enzyme
Compound
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Target Concepts:
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Query: EC:2.7.11.25 (
MEKK1
)
1,856
document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)
MEKK2
and
MEKK3
are mitogen-activated protein kinase kinase kinases (MAP3 kinases) of 70 and 71 kDa respectively that are markedly homologous (94%) in their kinase domains. Both
MEKK2
and
MEKK3
are able to activate the Jun kinase pathway in vivo. However, following routine immunoprecipitation in Triton X-100,
MEKK2
but not
MEKK3
is able to effectively phosphorylate both SEK-1 and MEK-1 and to undergo autophosphorylation. Unexpectedly, both
MEKK2
and
MEKK3
are functional in an in vitro kinase assay when cells are solubilized with the closely related detergent, NP-40. Given the high homology between these kinases, we set out to relate this differential sensitivity to Triton X-100 to differences in primary structure. A set of chimeric molecules were generated and the loss of activity in Triton X-100 mapped to kinase domain II/III and specifically to serine 390 of
MEKK3
and valine 384 of
MEKK2
, residues immediately N-terminal to the active site lysine. Mutation of serine 390 of
MEKK3
to a valine (as is found in
MEKK2
) conferred catalytic activity to
MEKK3
in Triton X-100 whereas the reciprocal alteration of valine 384 of
MEKK2
to a serine conferred lack of activity in Triton X-100 to
MEKK2
. Search of the protein database identified only three kinases,
MEKK3
, Pbs2p and Dd-
PKI
, with a serine or threonine at this site. The presence of a serine or threonine adjacent to the active site lysine in protein kinases is rare and, in
MEKK3
, results in detergent instability.
...
PMID:In vitro activity of MEKK2 and MEKK3 in detergents is a function of a valine to serine difference in the catalytic domain. 1134 2
Vasodilator-stimulated phosphoprotein (VASP) is a major substrate of protein kinase A (PKA). Here we described the novel mechanism of VASP phosphorylation via cAMP-independent PKA activation. We showed that in human umbilical vein endothelial cells (HUVECs) alpha-thrombin induced phosphorylation of VASP. Specific inhibition of Galpha13 protein by the RGS domain of a guanine nucleotide exchange factor, p115RhoGEF, inhibited thrombin-dependent phosphorylation of VASP. More importantly, Galpha13-induced VASP phosphorylation was dependent on activation of RhoA and
mitogen-activated protein kinase kinase kinase
,
MEKK1
, leading to the stimulation of the NF-kappaB signaling pathway. alpha-Thrombin-dependent VASP phosphorylation was inhibited by small interfering RNA-mediated knockdown of RhoA, whereas Galpha13-dependent VASP phosphorylation was inhibited by a specific RhoA inhibitor botulinum toxin C3 and by a dominant negative mutant of
MEKK1
. We determined that Galpha13-dependent VASP phosphorylation was also inhibited by specific PKA inhibitors,
PKI
and H-89. In addition, the expression of phosphorylation-deficient IkappaB and pretreatment with the proteasome inhibitor MG-132 abolished Galpha13- and alpha-thrombin-induced VASP phosphorylation. In summary, we have described a novel pathway of Galpha13-induced VASP phosphorylation that involves activation of RhoA and
MEKK1
, phosphorylation and degradation of IkappaB, release of PKA catalytic subunit from the complex with IkappaB and NF-kappaB, and subsequent phosphorylation of VASP.
...
PMID:A novel mechanism of G protein-dependent phosphorylation of vasodilator-stimulated phosphoprotein. 1604 15
