Department of Neurosurgery and Stanford Stroke Center, Stanford University, Stanford, CA, USA.
Neuroscience. 2012 May 3;209:155-60. doi: 10.1016/j.neuroscience.2012.02.005. Epub 2012 Feb 11.
The extracellular signal-regulated kinase (ERK) 1/2 protein requires a dual phosphorylation at conserved threonine and tyrosine residues to be fully activated under normal physiological conditions. Thus, ERK1/2 kinase activity is often defined by the quantity of phosphorylated kinase. However, this may not accurately represent its true activity under certain pathological conditions. We investigated whether ERK1/2 kinase activity is proportional to its phosphorylation state in a rat focal ischemia model with and without rapid ischemic preconditioning. We showed that phosphorylated-ERK1/2 protein levels were increased 2.6±0.07-fold, and ERK1/2 kinase activity was increased 10.6±1.9-fold in animals receiving ischemic preconditioning alone without test ischemia compared with sham group (P<0.05, n=6/group), suggesting that phosphorylated-ERK1/2 protein levels represent its kinase activity under these conditions. However, preconditioning plus test ischemia robustly blocked ERK1/2 kinase activity, whereas it increased phosphorylated-ERK1/2 protein levels beyond those receiving test ischemia alone, suggesting that phosphorylated-ERK1/2 protein levels were not representative of actual kinase activity in this pathological condition. In conclusion, protein phosphorylation levels of ERK1/2 do not always correspond to kinase activity, thus, measuring the true kinase activity is essential.
细胞外信号调节激酶 (ERK) 1/2 蛋白在正常生理条件下需要在保守的苏氨酸和酪氨酸残基上进行双重磷酸化才能被完全激活。因此，ERK1/2 激酶活性通常通过磷酸化激酶的数量来定义。然而，在某些病理条件下，这可能无法准确反映其真实活性。我们研究了在有或没有快速缺血预处理的大鼠局灶性缺血模型中，ERK1/2 激酶活性是否与其磷酸化状态成正比。我们表明，与假手术组相比，单独接受缺血预处理的动物中磷酸化-ERK1/2 蛋白水平增加了 2.6±0.07 倍，ERK1/2 激酶活性增加了 10.6±1.9 倍（P<0.05，n=6/组），表明在这些条件下磷酸化-ERK1/2 蛋白水平代表其激酶活性。然而，预处理加测试缺血强烈阻断了 ERK1/2 激酶活性，而它增加了磷酸化-ERK1/2 蛋白水平，超过了仅接受测试缺血的水平，表明在这种病理条件下，磷酸化-ERK1/2 蛋白水平不能代表实际的激酶活性。总之，ERK1/2 的蛋白磷酸化水平并不总是与激酶活性相对应，因此，测量真正的激酶活性是至关重要的。
