The phorbol ester-dependent activator of the mitogen-activated protein kinase p42mapk is a kinase with specificity for the threonine and tyrosine regulatory sites.
The phorbol ester-dependent activator of the mitogen-activated protein kinase p42mapk is a kinase with specificity for the threonine and tyrosine regulatory sites.
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丝裂原激活蛋白激酶 p42mapk 的佛波酯依赖性激活剂是一种对苏氨酸和酪氨酸调节位点具有特异性的激酶。
DOI：
10.1073/pnas.89.12.5221
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发表时间：
1992
影响因子：
11.1
通讯作者：
Sturgill,TW
中科院分区：
文献类型：
--
作者：
Rossomando,A;Wu,J;Weber,MJ;Sturgill,TW
丝裂原激活蛋白激酶（MAP 激酶）通过酪氨酸和苏氨酸双重磷酸化来响应各种刺激（包括佛波酯）而被激活。为了确定激活机制，使用重组野生型 42-kDa MAP 激酶 (p42mapk) 和 p42mapk 激酶缺陷突变体 (K52R) 来测定受刺激的 EL4.I12 小鼠胸腺瘤细胞中 p42mapk 的激活剂活性和 K52R 的激酶活性。佛波醇 12,13-二丁酸酯（10 分钟，650 nM）刺激 MAP 激酶激活剂的单峰，该激活剂在 pH 7.5 和 8.9 下从 Mono Q 与 K52R 激酶活性共洗脱。这两种活性均被丝氨酸/苏氨酸特异性磷酸酶 2A 灭活，但不被酪氨酸特异性磷酸酶 CD45 灭活。 K52R 的磷酸化特异发生在 Thr-183 和 Tyr-185 上，通过胰蛋白酶磷酸肽作图与合成标记磷酸肽进行比较来确定。这些发现表明佛波酯刺激的 MAP 激酶激酶可以通过苏氨酸和酪氨酸磷酸化激活 p42mapk，因此 p42mapk 不需要自磷酸化反应。
Mitogen-activated protein kinases (MAP kinases) are activated by dual tyrosine and threonine phosphorylations in response to various stimuli, including phorbol esters. To define the mechanism of activation, recombinant wild-type 42-kDa MAP kinase (p42mapk) and a kinase-defective mutant of p42mapk (K52R) were used to assay both activator activity for p42mapk and kinase activity toward K52R in stimulated EL4.I12 mouse thymoma cells. Phorbol 12,13-dibutyrate (10 min, 650 nM) stimulated a single peak of MAP kinase activator that was coeluted from Mono Q at pH 7.5 and 8.9 with K52R kinase activity. Both activities were inactivated by the serine/threonine-specific phosphatase 2A but not by the tyrosine-specific phosphatase CD45. Phosphorylation of K52R occurred specifically on Thr-183 and Tyr-185, as determined by tryptic phosphopeptide mapping in comparison with synthetic marker phosphopeptides. These findings indicate that phorbol ester-stimulated MAP kinase kinase can activate p42mapk by threonine and tyrosine phosphorylations, and that p42mapk thus does not require an autophosphorylation reaction.