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Ser-474 is the major target of insulin-mediated phosphorylation of protein kinase B beta in primary rat adipocytes.
Ludwiginstitutet för Cancerforskning.
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2002 (English)In: Cell Signal, ISSN 0898-6568, Vol. 14, no 2, 175-82 p.Article in journal (Refereed) Published
Abstract [en]

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.

Place, publisher, year, edition, pages
2002. Vol. 14, no 2, 175-82 p.
Keyword [en]
Adipocytes/drug effects/*enzymology, Animals, Cells; Cultured, Electrophoresis; Gel; Two-Dimensional, Enzyme Inhibitors/pharmacology, Insulin/*pharmacology, Intracellular Membranes/enzymology, Male, Oxazoles/pharmacology, Phosphoprotein Phosphatase/antagonists & inhibitors, Phosphorylation, Phosphoserine/*metabolism, Protein Transport, Protein-Serine-Threonine Kinases, Proto-Oncogene Proteins/*chemistry/*metabolism, Proto-Oncogene Proteins c-akt, Rats, Rats; Sprague-Dawley
URN: urn:nbn:se:uu:diva-10432PubMedID: 11781143OAI: oai:DiVA.org:uu-10432DiVA: diva2:38200
Available from: 2007-03-23 Created: 2007-03-23 Last updated: 2011-01-13

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