An insulin-stimulated ribosomal protein S6 kinase in 3T3-L1 cells.
A protein kinase that is stimulated from 2-10-fold by insulin and that phosphorylates ribosomal protein S6 has been characterized in 3T3-L1 cells. The detection of this activity in the 100,000 X g supernatant is facilitated by the presence of beta-glycerol phosphate or vanadate in the homogenization buffer. The activity has been purified 55-fold by chromatography on DEAE-cellulose and phosphocellulose. The resulting specific activity is 584 pmol/min/mg of protein. DEAE-cellulose chromatography followed by gel filtration on Ultrogel AcA54 or by glycerol gradient centrifugation suggests that the protein has a molecular mass of 60,000-70,000 daltons. Mg2+, and to a lesser extent Mn2+, will support phosphorylation of S6 by the activity. No proteins tested other than ribosomal protein S6 are phosphorylated. Based on its chromatographic properties and substrate specificity, the enzyme appears to be distinct from several other protein kinases that are known to phosphorylate ribosomal protein S6 in vitro. The complete characterization and purification of this enzyme may be essential to the elucidation of the mechanism of regulation of S6 phosphorylation by insulin.
D000818:Animals; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D005347:Fibroblasts / Q000201:enzymology; D005994:Glycerophosphates / Q000378:metabolism; D007328:Insulin / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D008970:Molecular Weight; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins / Q000378:metabolism*; D013379:Substrate Specificity; D014357:Trypsin / Q000378:metabolism
D005994:Glycerophosphates; D007328:Insulin; D007527:Isoenzymes; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; D008345:Manganese; D011494:Protein Kinases; D014357:Trypsin; D008274:Magnesium; C031463:beta-glycerophosphoric acid