Koontz S W, Jakubowski H, Goldman E
J Mol Biol. 1983 Aug 25;168(4):747-63. doi: 10.1016/s0022-2836(83)80073-4.
The effect of varying the concentration of Trp-tRNATrp in Escherichia coli infected with bacteriophage MS2 has been studied by varying the amount of exogenously added tryptophan (Trp) to cells bearing a mutation which results in a tryptophanyl-tRNA synthetase with a higher Km value for Trp. The phenotype of the mutant has been confirmed by measuring the level of tRNATrp which can be aminoacylated in vivo, and the mutant has also been shown to have elevated tRNATrp levels compared to wild-type. The growth of MS2 decreases continuously with decreasing Trp concentration (and hence, decreasing Trp-tRNATrp concentration). This appears to be due to reduced gene expression, since at later times in infection the amount of MS2 coat protein synthesized likewise decreases continuously with decreasing Trp concentration. However, there is little decrease in the amount of coat protein or replicase synthesized during the first few minutes after the Trp concentration shift. A continuous increase in the average polysome size distribution is seen as the Trp concentration is decreased. MS2 RNA synthesis also decreases continuously with decreasing Trp concentration, and is shut off in the absence of Trp. This does not seem to be due to ppGpp as these cells are functionally relaxed under these conditions, nor does it seem to be due to degradation of pre-existing template. Addition of chloramphenicol abolishes the effect of Trp concentration on RNA synthesis. The data are consistent with a model in which ribosomes pause at Trp codons in the absence of Trp-tRNATrp, while other ribosomes queue behind and continue to load onto the message. The reduction of RNA synthesis would then be a consequence of coupling to translation.
通过改变外源添加到携带一种突变的细胞中的色氨酸（Trp）的量，研究了在感染噬菌体MS2的大肠杆菌中改变色氨酸 - tRNA(^{Trp})浓度的影响，该突变导致色氨酰 - tRNA合成酶对Trp具有更高的Km值。通过测量体内可进行氨酰化的tRNA(^{Trp})水平，证实了该突变体的表型，并且还表明该突变体与野生型相比，tRNA(^{Trp})水平升高。随着Trp浓度的降低（因此，色氨酸 - tRNA(^{Trp})浓度降低），MS2的生长持续下降。这似乎是由于基因表达减少，因为在感染后期，随着Trp浓度降低，合成的MS2外壳蛋白的量同样持续下降。然而，在Trp浓度变化后的最初几分钟内，合成的外壳蛋白或复制酶的量几乎没有减少。随着Trp浓度降低，平均多核糖体大小分布持续增加。MS2 RNA合成也随着Trp浓度降低而持续下降，并且在没有Trp的情况下关闭。这似乎不是由于ppGpp，因为这些细胞在这些条件下功能松弛，也似乎不是由于预先存在的模板的降解。添加氯霉素消除了Trp浓度对RNA合成的影响。这些数据与一个模型一致，在该模型中，在没有色氨酸 - tRNA(^{Trp})的情况下，核糖体在Trp密码子处暂停，而其他核糖体在后面排队并继续加载到信使上。然后RNA合成的减少将是与翻译偶联的结果。
