Maki Jenny L, Degterev Alexei
Department of Biochemistry, School of Medicine, Tufts University, Boston, MA, USA.
Methods Mol Biol. 2013;1004:31-42. doi: 10.1007/978-1-62703-383-1_3.
Necroptosis is a novel form of regulated non-apoptotic cell death, which displays morphological features of necrosis. The kinase activity of receptor-interacting protein kinase 1 (RIP1) is a critical component in signaling for necroptosis. The development of assays to evaluate RIP1 kinase activity is important in the further development of existing and novel inhibitors of necroptosis. Here, we describe RIP1 protein expression and purification from mammalian and insect cells as well as two in vitro kinase assays to detect RIP1 kinase activity and inhibition.
坏死性凋亡是一种新型的受调控的非凋亡性细胞死亡形式，具有坏死的形态学特征。受体相互作用蛋白激酶1（RIP1）的激酶活性是坏死性凋亡信号传导的关键组成部分。开发评估RIP1激酶活性的检测方法对于现有和新型坏死性凋亡抑制剂的进一步开发至关重要。在此，我们描述了从哺乳动物和昆虫细胞中表达和纯化RIP1蛋白，以及两种用于检测RIP1激酶活性和抑制作用的体外激酶检测方法。
