Whelly S M, Baserga R
Biochim Biophys Acta. 1976 Mar 4;425(2):234-45. doi: 10.1016/0005-2787(76)90030-7.
A cell-free protein synthesis system employing ribosomes from WI-38 human diploid fibroblasts was developed and its optimum MgC12 and KC1 levels and pH value found. The rate at which ribosomes are able to incorporate radioactive leucine into proteins ([14C]leucine incorporation/10 min/100 mug rRNA) and the number of growing peptide chains [3H]puromycinpeptides formed/100 mug rRNA) was determined. When confluent monolayers of WI-38 cells were stimulated to proliferate by serum, a transient increase in the rate of peptide elongation by ribosomes was observed at 60 min after stimulation. This increase was not affected by the presence of actinomycin D (10 mug/ml) in the stimulating medium. A change in the relative amount of certain ribosome-associated proteins accompanied the increased elongation rate of peptide growth. The alteration in associated proteins could not be accounted for by an increased synthesis of protein. Finally, the early activation of ribosomes in stimulated WI-38 cells appears to result from the removal of an inhibitor(s) of ribosome function.
开发了一种利用WI-38人二倍体成纤维细胞核糖体的无细胞蛋白质合成系统，并确定了其最佳MgC12、KC1水平和pH值。测定了核糖体将放射性亮氨酸掺入蛋白质的速率（[14C]亮氨酸掺入量/10分钟/100微克rRNA）以及形成的正在生长的肽链数量（[3H]嘌呤霉素肽/100微克rRNA）。当用血清刺激汇合的WI-38细胞单层使其增殖时，在刺激后60分钟观察到核糖体肽链延伸速率短暂增加。这种增加不受刺激培养基中放线菌素D（10微克/毫升）存在的影响。肽生长延伸速率增加伴随着某些核糖体相关蛋白质相对量的变化。相关蛋白质的改变不能用蛋白质合成增加来解释。最后，受刺激的WI-38细胞中核糖体的早期激活似乎是由于核糖体功能抑制剂的去除。
