A cell free system from HeLa cells active in initiation of protein synthesis.
A cell free system from HeLa cells active in initiation of protein synthesis.
复制标题
来自 HeLa 细胞的无细胞系统，在蛋白质合成的起始过程中具有活性。
DOI：
10.1021/bi00695a015
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发表时间：
1975
期刊：
影响因子：
2.9
通讯作者：
C. Baglioni
中科院分区：
文献类型：
--
作者：
L. Weber;E. Feman;C. Baglioni
通过向用于均质化细胞的培养基中加入25-100 μ M氯化血红素，从HeLa细胞中获得了由内源性mRNA编程并在蛋白质合成起始中具有活性的无细胞系统。氯化血红素稳定了提取物的引发活性，否则即使在0 ℃下也会迅速衰减。氯化血红素在促进起始中的作用已经通过将提取物分级分离成核糖体和核糖体后上清液（S150）来检查。不含氯化血红素制备的提取物或由该不含氯化血红素制备的提取物获得的S150或由该提取物获得的S150抑制含氯化血红素的提取物的蛋白质合成约30%。从含有氯化血红素的提取物中制备的核糖体在蛋白质合成的起始中是活性的，而从不含氯化血红素的提取物中获得的核糖体显示很少或没有起始。这些结果表明，除了氯化血红素防止在S150的起始抑制剂的形成，并在同一时间保护与核糖体或核糖体亚基相关的起始因子的失活。添加2 mM GTP的HeLa提取物稳定的起始活性，虽然在较小的程度比氯化血红素。氯化血红素和GTP的作用不是相加的，这表明它们可能作用于相同的靶分子，尽管可能通过不同的机制。鉴于在兔网织红细胞无细胞系统中所作的类似观察，讨论了GTP的作用机制。
A cell free system programmed by endogenous mRNA and active in initiation of protein synthesis has been obtained from HeLa cells by adding 25-100 muM hemin to the medium used to homogenize the cells. Hemin stabilizes the initiation activity of the extract, which otherwise decays rapidly even at 0 degrees C. The role of hemin in promoting initiation has been examined by fractionating the extracts into ribosomes and postribosomal supernatant (S150). An extract prepared without hemin or the S150 obtained from this extract prepared without hemin or the S150 obtained from this extract inhibits protein synthesis of the extract containing hemin by about 30%. The ribosomes prepared from extracts containing hemin are active in initiation of protein synthesis, whereas the ribosomes obtained from the extracts prepared without hemin show little or no initiation. These results have suggested that addition of hemin prevents the formation of an inhibitor of initiation in the S150 and at the same time protects from inactivation an initiation factor associated with ribosomes or ribosomal subunits. Addition of 2 mM GTP to HeLa extracts stabilizes the initiation activity, though to a smaller degree than hemin. The effects of hemin and GTP are not additive, suggesting that they may act on the same target molecule, though possibly by different mechanisms. The mechanism of action of GTP is discussed in view of similar observations made in the rabbit reticulocyte cell free system.