SYNTHESIS OF LOW-MOLECULAR WEIGHT INHIBITOR OF PROTEIN-SYNTHESIS WITH ENZYME FROM INTERFERON-TREATED CELLS
SYNTHESIS OF LOW-MOLECULAR WEIGHT INHIBITOR OF PROTEIN-SYNTHESIS WITH ENZYME FROM INTERFERON-TREATED CELLS
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DOI：
10.1038/268537a0
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发表时间：
1977-01-01
期刊：
影响因子：
64.8
通讯作者：
KERR, IM
中科院分区：
文献类型：
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作者：
HOVANESSIAN, AG;BROWN, RE;KERR, IM
经抗病毒药物干扰素1处理的小鼠L细胞在无细胞系统中的蛋白质合成对双链核糖核酸(DsRNA)抑制的敏感性增强(参考文献2-4)。蛋白质合成的抑制依赖于与三磷酸腺苷和双链RNA的孵育，我们和其他人报道了干扰素处理的细胞5-8的提取物中存在双链RNA依赖的蛋白激酶活性(S)。病毒双链RNA介导的蛋白质合成抑制可能参与了病毒感染后的一系列事件，干扰素处理的细胞已经讨论过2，4。干扰素处理的细胞5-8的提取物的情况让人想起在兔网织红细胞裂解物中的情况，在这种情况下，蛋白激酶对起始因子的磷酸化涉及到在包括dsRNA9-12存在的各种条件下抑制蛋白质的合成。然而，在干扰素处理的L细胞系统中，除了蛋白激酶外，还在与三磷酸腺苷和双链RNA(REF)孵育时形成了一种热稳定的低分子量蛋白质合成抑制物(LMW抑制物)。6)。根据我们以前的工作6，LMW抑制物可能是dsRNA依赖的激酶的一种小的磷酸化的多肽产物。我们在这里表明，情况并非如此。此外，我们还证明了负责合成LMW抑制剂的酶将结合到固相载体上的dsRNA柱上。它可以在高盐中洗脱，但在这种形式下相对不稳定。我们利用高纯度的酶在其稳定的柱结合状态下合成并放射性标记了LMW抑制剂。本文报道了该抑制剂的合成、部分纯化及其对L细胞和兔网织红细胞无细胞系统蛋白质合成的抑制作用。更详细的缓蚀剂特征描述见所附文件13。
PROTEIN synthesis in cell-free systems from mouse L cells pretreated with the antiviral agent interferon1shows an enhanced sensitivity to inhibition by double-stranded RNA (dsRNA) (refs 2–4). The inhibition of protein synthesis is dependent on incubation with ATP as well as dsRNA and we and others have reported a dsRNA-dependent protein kinase activity(s) in extracts from interferon-treated cells5–8. The possible involvement of a viral dsRNA-mediated inhibition of protein synthesis in the sequence of events following virus infection in intact, interferon-treated cells has been discussed previously2,4. The situation with extracts from interferon-treated cells5–8is reminiscent of that in rabbit reticulocyte lysates in which phosphorylation of an initiation factor by a protein kinase has been implicated in the inhibition of protein synthesis in a variety of conditions including the presence of dsRNA9–12. In the interferon-treated L-cell system, however, in addition to the kinase, a heat-stable, low molecular weight inhibitor (LMW inhibitor) of protein synthesis is formed on incubation with ATP and dsRNA (ref. 6). It remained possible from our previous work6, that the LMW inhibitor might be a small phosphorylated peptide product of the dsRNA-dependent kinase. We show here that this is not the case. In addition, we show that the enzyme responsible for the synthesis of the LMW inhibitor will bind to a column of dsRNA attached to a solid support. It can be eluted in high salt but is relatively unstable in this form. We have used the highly-purified enzyme in its stable column-bound state to synthesise and radioactively label the LMW inhibitor. This synthesis, the partial purification of the inhibitor and its activity in the inhibition of protein synthesis in cell-free systems from L cells and rabbit reticulocytes, are reported here. A more detailed characterisation of the inhibitor is described in the accompanying paper13.