RNA-DEPENDENT RNA-POLYMERASE ACTIVITY IN CORONAVIRUS-INFECTED CELLS
RNA-DEPENDENT RNA-POLYMERASE ACTIVITY IN CORONAVIRUS-INFECTED CELLS
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DOI：
10.1128/jvi.42.1.153-164.1982
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发表时间：
1982-01-01
影响因子：
5.4
通讯作者：
BRIAN, DA
中科院分区：
文献类型：
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作者：
DENNIS, DE;BRIAN, DA
在内源模板存在下，在感染猪传染性胃肠炎病毒的猪细胞的细胞质中发现了将[3H]UMP掺入酸可沉淀材料中的酶活性。在未感染的对照细胞中没有发现这种活性，在纯化的病毒中也没有发现这种活性。该活性与受感染细胞的线粒体部分相关，表明该酶是膜结合的。该活性需要除 [3H]UTP 之外的所有 3 种核糖核苷三磷酸的存在，并且它不受放线菌素 D 抑制。加热的产物被 RNase 消化，但不被 DNase 消化。酶活性需要Mg2+，其最佳浓度为.apprx。 5毫米。通过电泳将体外产物的大小与体内合成的病毒特异性RNA的大小进行比较，以确认聚合酶活性的病毒特异性。来自感染细胞的病毒特异性RNA由10种单链多聚腺苷酸RNA组成，MW为6.8倍。 106、6.2 倍。 106, 3.15 倍。 106、1.40 倍。 106, 1.05 倍。 106, 0.94 倍。 106, 0.66 倍。 106, 0.39 倍。 106, 0.34 倍。 106 和 0.24 倍。 106.体外合成的RNA由MW明显高于基因组RNA的高MW种类和与1.40倍的种类电泳共迁移的2个单链种类组成。 106 和 0.66 倍。 106 MW 体内制造。
An enzymatic activity which incorporates [3H]UMP into acid-precipitable material in the presence of endogenous template was found in the cytoplasm of porcine cells infected with the transmissible gastroenteritis virus of swine. This activity was not found in uninfected control cells, nor was it found in purified virus. The activity was associated with the mitochondrial fraction of infected cells, suggesting that the enzyme is membrane bound. The activity required the presence of all 3 ribonucleoside triphosphates in addition to [3H]UTP, and it was not inhibited by actinomycin D. The heated product was digested by RNase but not by DNase. Mg2+ was required for enzymatic activity, and its optimal concentration was .apprx. 5 mM. The size of the in vitro products was compared by electrophoresis with that of in vivo-synthesized virus-specified RNA to confirm the viral specificity of the polymerase activity. Virus-specified RNA from infected cells consisted of 10 species of single-stranded, polyadenylated RNA with MW of 6.8 .times. 106, 6.2 .times. 106, 3.15 .times. 106, 1.40 .times. 106, 1.05 .times. 106, 0.94 .times. 106, 0.66 .times. 106, 0.39 .times. 106, 0.34 .times. 106 and 0.24 .times. 106. In vitro-synthesized RNA consisted of a high MW species of apparently higher MW than genomic RNA, and 2 single-stranded species that electrophoretically comigrated with the species of 1.40 .times. 106 and 0.66 .times. 106 MW made in vivo.