Rojano-Nisimura Alejandra M, Haning Katie, Janovsky Justin, Vasquez Kevin A, Thompson Jeffrey P, Contreras Lydia M
Institute for Cellular and Molecular Biology , The University of Texas at Austin , 2500 Speedway Stop A4800 , Austin , Texas 78712 , United States.
McKetta Department of Chemical Engineering , The University of Texas at Austin , 200 E. Dean Keeton Street Stop C0400 , Austin , Texas 78712 , United States.
ACS Synth Biol. 2020 Feb 21;9(2):329-342. doi: 10.1021/acssynbio.9b00344. Epub 2019 Dec 10.
An intriguing aspect of protein synthesis is how cotranslational events are managed inside the cell. In this study, we developed an bimolecular fluorescence complementation assay coupled to SecM stalling (BiFC-SecM) to study how codon usage influences the interactions of ribosome-associating factors that occur cotranslationally. We profiled ribosomal associations of a number of proteins, and observed differential association of chaperone proteins TF, DnaK, GroEL, and translocation factor Ffh as a result of introducing synonymous codon substitutions that change the affinity of the translating sequence to the ribosomal anti-Shine-Dalgarno (aSD) sequence. The use of pausing sequences within proteins regulates their transit within the translating ribosome. Our results indicate that the dynamics between cellular factors and the new polypeptide chain are affected by how codon composition is designed. Furthermore, associating factors may play a role in processes including protein quality control (folding and degradation) and cellular respiration.
蛋白质合成中一个有趣的方面是细胞内共翻译事件是如何被管理的。在本研究中，我们开发了一种与SecM停滞相结合的双分子荧光互补分析方法（BiFC-SecM），以研究密码子使用如何影响共翻译过程中核糖体相关因子的相互作用。我们分析了多种蛋白质的核糖体关联情况，并观察到由于引入同义密码子替换改变了翻译序列与核糖体反Shine-Dalgarno（aSD）序列的亲和力，伴侣蛋白TF、DnaK、GroEL和转运因子Ffh出现了不同的关联。蛋白质内暂停序列的使用调节了它们在翻译核糖体中的转运。我们的结果表明，细胞因子与新多肽链之间的动态关系受密码子组成设计方式的影响。此外，相关因子可能在包括蛋白质质量控制（折叠和降解）及细胞呼吸等过程中发挥作用。
