THE BINDING OF DOUBLE-STRANDED-RNA AND ADENOVIRUS VAI RNA TO THE INTERFERON-INDUCED PROTEIN-KINASE
THE BINDING OF DOUBLE-STRANDED-RNA AND ADENOVIRUS VAI RNA TO THE INTERFERON-INDUCED PROTEIN-KINASE
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DOI：
10.1111/j.1432-1033.1989.tb14485.x
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发表时间：
1989-01-02
期刊：
影响因子：
--
通讯作者：
HOVANESSIAN, AG
中科院分区：
文献类型：
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作者：
GALABRU, J;KATZE, MG;HOVANESSIAN, AG
来自依赖于双链（ds）RNA的人细胞的蛋白激酶是68-kDa蛋白（p68激酶），其水平在用干扰素处理的细胞中显著增强。当被低浓度的dsRNA激活时，p68激酶被磷酸化，从而催化蛋白质合成起始因子eIF 2的磷酸化。在此，我们使用特异性单克隆抗体将p68激酶纯化至同质，以研究其结合dsRNA的能力，poly（I）.聚（C）。我们的研究表明，p68激酶具有高亲和力和低亲和力结合位点：高亲和力结合位点负责激活和低亲和力结合位点抑制激酶活性。这与p68激酶的自磷酸化在低浓度dsRNA下发生而高浓度dsRNA抑制其自磷酸化的事实雅阁。我们还研究了腺病毒VAI RNA与纯化的p68激酶的结合，并发现这种结合的亲和力低于poly（I）的亲和力。聚（C）。我们表明，VAI RNA可以激活或抑制p68激酶的自磷酸化的剂量依赖性的方式，即激活在≤ 200 μ mol/L。在> 1 μ g/ml的VAI RNA下抑制或在> 1 μ g/ml时抑制。尽管VAI RNA的结合亲和力较低，但其不能被poly（I）置换。聚（C）或呼肠孤病毒dsRNA。这些数据证实了我们以前的结果，说明VAI RNA可以结合p68激酶，并导致其失活不可逆。
The protein kinase from human cells dependent on double-stranded (ds) RNA is a 68-kDa protein (p68 kinase), the level of which is enhanced significantly in cells treated with interferon. When activated by low concentrations of dsRNA, the p68 kinase becomes phosphorylated and thereby catalyzes the phosphorylation of the protein-synthesis initiation factor, eIF2. Here, we have purified the p68 kinase to homogeneity using a specific monoclonal antibody to investigate its capacity to bind dsRNA, poly(I) .cntdot. poly(C). Our study suggest that p68 kinase has high- and low-affinity binding sites: the high-affinity binding site is responsible for the activation and the low-affinity binding site for the inhibition of kinase activity. This is in accord with the fact that autophosphorylation of p68 kinase occurs at low concentrations of dsRNA whereas high concentrations of dsRNA inhibit its autophosphorylation. We have also investigated the binding of adenoviral VAI RNA to the purified p68 kinase and have found that the affinity of this binding is lower than that of poly(I) .cntdot. poly(C). We show that VAI RNA can activate or inhibit autophosphorylation of p68 kinase in a dose-dependent manner, i.e. activation at .ltoreq. 1 .mu.g/ml or inhibition at > 1 .mu.g/ml of VAI RNA. In spite of its lower affinity of binding, VAI RNA cannot be displaced by poly(I) .cntdot. poly(C) or reovirus dsRNA. These data confirm our previous results to illustrate that VAI RNA can bind p68 kinase and cause its inactivation irreversably.