Phosphorylation of GRK2 by protein kinase C abolishes its inhibition
by calmodulin
C. Krasel, S. Dammeier, R. Winstel, J. Brockmann, H. Mischak, and M. Lohse. J Biol Chem, 276 (3):
1911-5(January 2001)Krasel, C Dammeier, S Winstel, R Brockmann, J Mischak, H Lohse, M
J Research Support, Non-U.S. Gov't United States The Journal of biological
chemistry J Biol Chem. 2001 Jan 19;276(3):1911-5. Epub 2000 Oct 19..
Abstract
G-protein-coupled receptor kinases (GRKs) are important regulators
of G-protein-coupled receptor function. Two members of this family
L, GRK2 and GRK5 L, have been shown to be substrates for protein
kinase C (PKC). Whereas PKC-mediated phosphorylation results in inhibition
of GRK5, it increases the activity of GRK2 toward its substrates
probably through increased affinity for receptor-containing membranes.
We show here that this increase in activity may be caused by relieving
a tonic inhibition of GRK2 by calmodulin. In vitro, GRK2 was preferentially
phosphorylated by PKC isoforms alpha, gamma, and delta. Two-dimensional
peptide mapping of PKCalpha-phosphorylated GRK2 showed a single site
of phosphorylation, which was identified as serine 29 by HPLC-MS.
A S29A mutant of GRK2 was not phosphorylated by PKC in vitro and
showed no phorbol ester-stimulated phosphorylation when transfected
into human embryonic kidney (HEK)293 cells. Serine 29 is located
in the calmodulin-binding region of GRK2, and binding of calmodulin
to GRK2 results in inhibition of kinase activity. This inhibition
was almost completely abolished in vitro when GRK2 was phosphorylated
by PKC. These data suggest that calmodulin may be an inhibitor of
GRK2 whose effects can be abolished with PKC-mediated phosphorylation
of GRK2.
Krasel, C Dammeier, S Winstel, R Brockmann, J Mischak, H Lohse, M
J Research Support, Non-U.S. Gov't United States The Journal of biological
chemistry J Biol Chem. 2001 Jan 19;276(3):1911-5. Epub 2000 Oct 19.
%0 Journal Article
%1 Krasel2001
%A Krasel, C.
%A Dammeier, S.
%A Winstel, R.
%A Brockmann, J.
%A Mischak, H.
%A Lohse, M. J.
%D 2001
%J J Biol Chem
%K & 2 AMP-Dependent Animals C/*metabolism Calmodulin/*metabolism Cell Chromatography, Cyclic G-Protein-Coupled High Humans Isoenzymes/*metabolism Kinase Kinases Kinases/antagonists Line Liquid Mass Mutagenesis, Phosphorylation Pressure Protein Receptor Serine/metabolism Site-Directed Specificity Spectrometry Substrate beta-Adrenergic inhibitors/genetics/*metabolism
%N 3
%P 1911-5
%T Phosphorylation of GRK2 by protein kinase C abolishes its inhibition
by calmodulin
%U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11042191
%V 276
%X G-protein-coupled receptor kinases (GRKs) are important regulators
of G-protein-coupled receptor function. Two members of this family
L, GRK2 and GRK5 L, have been shown to be substrates for protein
kinase C (PKC). Whereas PKC-mediated phosphorylation results in inhibition
of GRK5, it increases the activity of GRK2 toward its substrates
probably through increased affinity for receptor-containing membranes.
We show here that this increase in activity may be caused by relieving
a tonic inhibition of GRK2 by calmodulin. In vitro, GRK2 was preferentially
phosphorylated by PKC isoforms alpha, gamma, and delta. Two-dimensional
peptide mapping of PKCalpha-phosphorylated GRK2 showed a single site
of phosphorylation, which was identified as serine 29 by HPLC-MS.
A S29A mutant of GRK2 was not phosphorylated by PKC in vitro and
showed no phorbol ester-stimulated phosphorylation when transfected
into human embryonic kidney (HEK)293 cells. Serine 29 is located
in the calmodulin-binding region of GRK2, and binding of calmodulin
to GRK2 results in inhibition of kinase activity. This inhibition
was almost completely abolished in vitro when GRK2 was phosphorylated
by PKC. These data suggest that calmodulin may be an inhibitor of
GRK2 whose effects can be abolished with PKC-mediated phosphorylation
of GRK2.
@article{Krasel2001,
abstract = {G-protein-coupled receptor kinases (GRKs) are important regulators
of G-protein-coupled receptor function. Two members of this family
L, GRK2 and GRK5 L, have been shown to be substrates for protein
kinase C (PKC). Whereas PKC-mediated phosphorylation results in inhibition
of GRK5, it increases the activity of GRK2 toward its substrates
probably through increased affinity for receptor-containing membranes.
We show here that this increase in activity may be caused by relieving
a tonic inhibition of GRK2 by calmodulin. In vitro, GRK2 was preferentially
phosphorylated by PKC isoforms alpha, gamma, and delta. Two-dimensional
peptide mapping of PKCalpha-phosphorylated GRK2 showed a single site
of phosphorylation, which was identified as serine 29 by HPLC-MS.
A S29A mutant of GRK2 was not phosphorylated by PKC in vitro and
showed no phorbol ester-stimulated phosphorylation when transfected
into human embryonic kidney (HEK)293 cells. Serine 29 is located
in the calmodulin-binding region of GRK2, and binding of calmodulin
to GRK2 results in inhibition of kinase activity. This inhibition
was almost completely abolished in vitro when GRK2 was phosphorylated
by PKC. These data suggest that calmodulin may be an inhibitor of
GRK2 whose effects can be abolished with PKC-mediated phosphorylation
of GRK2.},
added-at = {2010-12-14T18:12:02.000+0100},
author = {Krasel, C. and Dammeier, S. and Winstel, R. and Brockmann, J. and Mischak, H. and Lohse, M. J.},
biburl = {https://www.bibsonomy.org/bibtex/24bbfe89926ac1930614321ce4255000e/pharmawuerz},
endnotereftype = {Journal Article},
interhash = {6941f5c0710e00b6b740b617ddf10e62},
intrahash = {4bbfe89926ac1930614321ce4255000e},
issn = {0021-9258 (Print) 0021-9258 (Linking)},
journal = {J Biol Chem},
keywords = {& 2 AMP-Dependent Animals C/*metabolism Calmodulin/*metabolism Cell Chromatography, Cyclic G-Protein-Coupled High Humans Isoenzymes/*metabolism Kinase Kinases Kinases/antagonists Line Liquid Mass Mutagenesis, Phosphorylation Pressure Protein Receptor Serine/metabolism Site-Directed Specificity Spectrometry Substrate beta-Adrenergic inhibitors/genetics/*metabolism},
month = {Jan 19},
note = {Krasel, C Dammeier, S Winstel, R Brockmann, J Mischak, H Lohse, M
J Research Support, Non-U.S. Gov't United States The Journal of biological
chemistry J Biol Chem. 2001 Jan 19;276(3):1911-5. Epub 2000 Oct 19.},
number = 3,
pages = {1911-5},
shorttitle = {Phosphorylation of GRK2 by protein kinase C abolishes its inhibition
by calmodulin},
timestamp = {2010-12-14T18:22:02.000+0100},
title = {Phosphorylation of GRK2 by protein kinase C abolishes its inhibition
by calmodulin},
url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11042191},
volume = 276,
year = 2001
}