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Ser-474 is the major target of insulin-mediated phosphorylation of protein kinase B beta in primary rat adipocytes.

Göransson, Olga LU ; Resjö, Svante LU ; Rönnstrand, Lars; Manganiello, Vincent and Degerman, Eva LU (2002) In Cellular Signalling 14(2). p.175-182
Abstract
The mechanism of activation for protein kinase B (PKB), an important target for insulin signaling, has been scarcely investigated in primary cells. In this study, we have characterized the insulin-induced phosphorylation and activation of PKB beta in primary rat adipocytes. Insulin stimulation resulted in a translocation of PKB beta from cytosol to membranes, and phosphorylation and activation of PKB beta. Phosphoamino acid analysis and phosphopeptide mapping demonstrated that the phosphorylation occurred mainly on serines, also when using calyculin A, and that these were localized within one major phosphopeptide. Radiosequencing showed that the radioactivity was released in Cycle No. 7. In addition, the peptide was specifically... (More)
The mechanism of activation for protein kinase B (PKB), an important target for insulin signaling, has been scarcely investigated in primary cells. In this study, we have characterized the insulin-induced phosphorylation and activation of PKB beta in primary rat adipocytes. Insulin stimulation resulted in a translocation of PKB beta from cytosol to membranes, and phosphorylation and activation of PKB beta. Phosphoamino acid analysis and phosphopeptide mapping demonstrated that the phosphorylation occurred mainly on serines, also when using calyculin A, and that these were localized within one major phosphopeptide. Radiosequencing showed that the radioactivity was released in Cycle No. 7. In addition, the peptide was specifically immunoprecipitated from a tryptic digest of PKB beta using the anti-phospho-PKB (Ser-473) antibody. Taken together, these results show that rat adipocyte PKB beta mainly is phosphorylated on Ser-474 in response to insulin stimulation, in contrast to previous studies in human embryonic kidney (HEK) 293 cells demonstrating, in addition, phosphorylation of Thr-309. (Less)
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keywords
Intracellular Membranes : enzymology, Oxazoles : pharmacology, Phosphoprotein Phosphatase : antagonists & inhibitors, Phosphorylation, Phosphoserine : metabolism, Protein Transport, Proto-Oncogene Proteins : chemistry : metabolism, Rats, Support Non-U.S. Gov't, Rats Sprague-Dawley, Insulin : pharmacology, Male, Enzyme Inhibitors : pharmacology, Cells Cultured, Electrophoresis Gel Two-Dimensional, Animal, Adipocytes : drug effects : enzymology
in
Cellular Signalling
volume
14
issue
2
pages
175 - 182
publisher
Elsevier
external identifiers
  • wos:000173433000012
  • scopus:0036144090
ISSN
1873-3913
DOI
10.1016/S0898-6568(01)00242-X
language
English
LU publication?
yes
id
eb2ab288-2c76-47b1-a2b0-f5341b72b2a4 (old id 106800)
alternative location
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11781143&dopt=Abstract
date added to LUP
2007-07-12 14:58:58
date last changed
2017-01-01 04:57:22
@article{eb2ab288-2c76-47b1-a2b0-f5341b72b2a4,
  abstract     = {The mechanism of activation for protein kinase B (PKB), an important target for insulin signaling, has been scarcely investigated in primary cells. In this study, we have characterized the insulin-induced phosphorylation and activation of PKB beta in primary rat adipocytes. Insulin stimulation resulted in a translocation of PKB beta from cytosol to membranes, and phosphorylation and activation of PKB beta. Phosphoamino acid analysis and phosphopeptide mapping demonstrated that the phosphorylation occurred mainly on serines, also when using calyculin A, and that these were localized within one major phosphopeptide. Radiosequencing showed that the radioactivity was released in Cycle No. 7. In addition, the peptide was specifically immunoprecipitated from a tryptic digest of PKB beta using the anti-phospho-PKB (Ser-473) antibody. Taken together, these results show that rat adipocyte PKB beta mainly is phosphorylated on Ser-474 in response to insulin stimulation, in contrast to previous studies in human embryonic kidney (HEK) 293 cells demonstrating, in addition, phosphorylation of Thr-309.},
  author       = {Göransson, Olga and Resjö, Svante and Rönnstrand, Lars and Manganiello, Vincent and Degerman, Eva},
  issn         = {1873-3913},
  keyword      = {Intracellular Membranes : enzymology,Oxazoles : pharmacology,Phosphoprotein Phosphatase : antagonists & inhibitors,Phosphorylation,Phosphoserine : metabolism,Protein Transport,Proto-Oncogene Proteins : chemistry : metabolism,Rats,Support Non-U.S. Gov't,Rats Sprague-Dawley,Insulin : pharmacology,Male,Enzyme Inhibitors : pharmacology,Cells Cultured,Electrophoresis Gel Two-Dimensional,Animal,Adipocytes : drug effects : enzymology},
  language     = {eng},
  number       = {2},
  pages        = {175--182},
  publisher    = {Elsevier},
  series       = {Cellular Signalling},
  title        = {Ser-474 is the major target of insulin-mediated phosphorylation of protein kinase B beta in primary rat adipocytes.},
  url          = {http://dx.doi.org/10.1016/S0898-6568(01)00242-X},
  volume       = {14},
  year         = {2002},
}