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SmpB-tmRNA介导的反式翻译,是细菌中普遍存在的一种主要核糖体拯救机制,对细菌的生存和增殖都具有重要影响。为了探明反式翻译系统在人-鱼共患病原菌维氏气单胞菌(Aeromonas veronii)中的作用机制,本研究基于维氏气单胞菌tmRNA的二级结构预测,将tmRNA标记肽的后5个氨基酸残基的密码子以及1个终止密码子突变为组氨酸密码子,从而构建tmRNA突变体。生长曲线测定结果表明,该tmRNA突变体能够回补tmRNA缺失导致的细菌生长缺陷;同时,免疫印迹实验确证该突变体能够成功地将反式翻译拯救的蛋白底物标记上组氨酸标签,表明其能够行使tmRNA的正常功能。本研究构建的tmRNA突变体为后续分离和纯化tmRNA-SmpB介导的反式翻译底物,进而研究反式翻译系统在细菌中的作用机制提供了理论基础。
Abstract：The trans-translation mediated by SmpB-tmRNA is a major ribosome rescue mechanism prevalent in bacteria, and has significant influences on bacterial survival and proliferation. In order to investigate the action mechanism of trans-translation system in Human-fish pathogen Aeromonas veronii, a mutant of tmRNA was constructed by mutating the codon of the last five amino acid residues of tmRNA-labeled peptide and one termination codon into histidine codon based on the prediction of the secondary structure of tmRNA in Aeromonas veronii. The results of growth curve determination showed that the tmRNA mutant could compensate for the defect of bacterial growth caused by the deletion of tmRNA. At the same time, Western blotting confirmed that the protein substrates rescued by trans-translation were successfully labeled with histidine tags by the mutant, indicating that the tmRNA mutant normally functioned as that of tmRNA. The tmRNA mutant constructed in this study would provided a theoretical basis for the subsequent isolation and purification of trans-translation substrates mediated by tm RNA-Smp B,further study of trans-translation mechanism in bacteria.
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基本信息:
DOI：10.13417/j.gab.038.001636
中图分类号:Q933
引用信息:
[1]彭钼植,唐燕琼,唐鸿倩,等.维氏气单胞菌tmRNA突变体构建及其用于反式翻译底物的鉴定[J].基因组学与应用生物学,2019,38(04):1636-1642.DOI:10.13417/j.gab.038.001636.
基金信息:
科技部国际合作专项(2015DFR31060);; 海南省重点研发计划(ZDYF2017020);; 海南省自然科学基金(317015);; 国家自然科学基金项目(31360261;31560021;31772887);; 海南大学青年基金资助项目(HDKYXJ201712)共同资助
2018-01-08
2018-01-08
2018-01-08