Purification and properties of extracellular signal-regulated kinase 1, an insulin-stimulated microtubule-associated protein 2 kinase.
In rat 1 fibroblasts, insulin has little or no stimulatory effect on the activities of either MAP2 protein kinase or ribosomal protein S6 kinase. In contrast, in rat 1 cells that overexpress the normal human insulin receptor (rat 1 HIRc B; McClain et al. (1987) J. Biol. Chem. 262, 14663-14671), insulin activates both MAP2 and S6 kinase activities close to 5-fold. A MAP2 kinase has been purified from insulin-treated rat 1 HIRc B cells over 6300-fold by chromatography on Q-Sepharose, phenyl-Sepharose, S-Sepharose, phosphocellulose, QAE-Sepharose, UltrogelAcA54, DEAE-cellulose, and a second Q-Sepharose. Its specific activity is approximately 0.8-1 mumol.min-1.mg-1 with MAP2 and 3 mumol.min-1.mg-1 with myelin basic protein. The enzyme preparation contains one major band of Mr = 43,000 upon SDS-polyacrylamide gel electrophoresis, which is immunoblotted by antibodies to phosphotyrosine. A sequence from the 43-kDa band led to the isolation of a cDNA encoding the enzyme, which we have named ERK1 for extracellular signal-regulated kinase (Boulton et al. (1990) Science 249, 64-67).
Boulton
T G
TG
Department of Pharmacology, University of Texas Southwestern Graduate School of Biomedical Sciences, Dallas 75235-9041.
Gregory
J S
JS
Cobb
M H
MH
D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000201:enzymology; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002460:Cell Line; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D005347:Fibroblasts / Q000201:enzymology; D015151:Immunoblotting; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011494:Protein Kinases / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D013379:Substrate Specificity
D007328:Insulin; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases