Department of Biophysics and Physical Biochemistry, University of Regensburg, Postfach, D-93040 Regensburg, Federal Republic of Germany.
J Biotechnol. 2010 Oct 1;150(1):44-50. doi: 10.1016/j.jbiotec.2010.07.017. Epub 2010 Jul 16.
Cell-free production of proteins is a rapid developing technology for many applications in proteomic sciences. Although great progress was made in the last years, in vitro protein synthesis is still less efficient than in vivo protein synthesis. For Escherichia coli based systems, all factors needed for cell-free protein synthesis are established, as shown in the reconstituted PURE system. Here we report the influence of additional translation factors, aminoacyl-tRNA synthetases and translational active ribosomes on cell-free protein synthesis of E. coli S30 extracts, indicating that none of these factors is a limiting factor for the protein synthesis. Furthermore, we show that varying the ratio of ribosomes including associated factors to other factors present in the extracts could not increase the yield of protein synthesized. In summary our results provide strong evidence for an optimal reconstitution of the translation machinery in E. coli S30 extracts.
无细胞蛋白质生产是蛋白质组学科学中许多应用的快速发展技术。尽管近年来取得了巨大进展，但体外蛋白质合成的效率仍低于体内蛋白质合成。对于基于大肠杆菌的系统，已建立了无细胞蛋白质合成所需的所有因素，如重组 PURE 系统所示。在这里，我们报告了额外的翻译因子、氨酰-tRNA 合成酶和翻译活性核糖体对大肠杆菌 S30 提取物无细胞蛋白质合成的影响，表明这些因素都不是蛋白质合成的限制因素。此外，我们还表明，改变核糖体（包括相关因子）与提取物中其他因子的比例并不能提高合成蛋白质的产量。总之，我们的结果为大肠杆菌 S30 提取物中转译机制的最佳重建提供了有力证据。
