Ray L B, Sturgill T W
Proc Natl Acad Sci U S A. 1987 Mar;84(6):1502-6. doi: 10.1073/pnas.84.6.1502.
Insulin treatment (Kact, 5 X 10(-9) M) of serum-starved 3T3-L1 adipocytes stimulates a soluble serine/threonine kinase that catalyzes phosphorylation of microtubule-associated protein 2 (MAP-2) in vitro. Maximal activation of MAP-2 kinase activity by 80 nM insulin was observed after 10 min of hormonal stimulation, prior to maximal stimulation of S6 kinase activity (20 min). The insulin-stimulatable MAP-2 kinase activity is not adsorbed to phosphocellulose, whereas the principal S6 kinase activity is retained and elutes at approximately 0.5 M NaCl. The insulin-stimulatable MAP-2 kinase is less stable during incubation at 30 degrees C than S6 kinase activity. Inclusion of phosphatase inhibitors decreases the rate at which the stimulated MAP-2 kinase activity is lost from extract supernatants incubated at 30 degrees C. p-Nitrophenyl phosphate is more effective than DL-phosphotyrosine, whereas DL-phosphoserine is without effect at the concentration used (40 mM). The difference in MAP-2 kinase activity in extract supernatants from control and insulin-treated cells is also preserved after rapid chromatography on Sephadex G-25. These results show that a soluble serine/threonine kinase is rapidly activated by insulin, possibly by phosphorylation of either the kinase itself or an interacting modulator.
用胰岛素（浓度为5×10⁻⁹ M）处理血清饥饿的3T3-L1脂肪细胞，可刺激一种可溶性丝氨酸/苏氨酸激酶，该激酶在体外催化微管相关蛋白2（MAP-2）的磷酸化。在激素刺激10分钟后观察到80 nM胰岛素对MAP-2激酶活性的最大激活，这早于S6激酶活性的最大刺激（20分钟）。胰岛素可刺激的MAP-2激酶活性不被磷酸纤维素吸附，而主要的S6激酶活性则被保留，并在约0.5 M NaCl浓度下洗脱。在30℃孵育期间，胰岛素可刺激的MAP-2激酶比S6激酶活性更不稳定。加入磷酸酶抑制剂可降低在30℃孵育的提取物上清液中受刺激的MAP-2激酶活性丧失的速率。对硝基苯磷酸比DL-磷酸酪氨酸更有效，而DL-磷酸丝氨酸在所使用的浓度（40 mM）下没有作用。在Sephadex G-25上快速层析后，对照细胞和胰岛素处理细胞的提取物上清液中MAP-2激酶活性的差异也得以保留。这些结果表明，一种可溶性丝氨酸/苏氨酸激酶可被胰岛素迅速激活，可能是通过激酶自身或相互作用的调节因子的磷酸化作用。
