Takai K, Takaku H, Yokoyama S
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Japan.
Nucleic Acids Res. 1996 Aug 1;24(15):2894-9. doi: 10.1093/nar/24.15.2894.
Unmodified tRNA molecules are useful for many purposes in cell-free protein biosynthesis, but there is little information about how the lack of tRNA post-transcriptional modifications affects the coding specificity for synonymous codons. In the present study, we prepared an unmodified form of Escherichia coli tRNA1Ser, which originally has the cmo5UGA anticodon (cmo5U = uridine 5-oxyacetic acid) and recognizes the UCU, UCA and UCG codons. The codon specificity of the unmodified tRNA was tested in a cell-free protein synthesis directed by designed mRNAs under competition conditions with the parent tRNA1Ser. It was found that the unmodified tRNA with the UGA anti-codon recognizes the UCA codon nearly as efficiently as the modified tRNA. The unmodified tRNA recognized the UCU codon with low, but detectable efficiency, whereas no recognition of the UCC and UCG codons was detected. Therefore, the absence of modifications makes this tRNA more specific to the UCA codon by remarkably reducing the efficiencies of wobble reading of other synonymous codons, without a significant decrease in the UCA reading efficiency.
未修饰的tRNA分子在无细胞蛋白质生物合成中有多种用途，但关于tRNA转录后修饰的缺失如何影响同义密码子的编码特异性，相关信息较少。在本研究中，我们制备了未修饰形式的大肠杆菌tRNA1Ser，它原本具有cmo5UGA反密码子（cmo5U = 5-氧乙酸尿苷），并识别UCU、UCA和UCG密码子。在与亲本tRNA1Ser竞争的条件下，在由设计的mRNA指导的无细胞蛋白质合成中测试了未修饰tRNA的密码子特异性。结果发现，带有UGA反密码子的未修饰tRNA识别UCA密码子的效率几乎与修饰后的tRNA相同。未修饰的tRNA以较低但可检测到的效率识别UCU密码子，而未检测到对UCC和UCG密码子的识别。因此，修饰的缺失通过显著降低其他同义密码子摆动阅读的效率，使这种tRNA对UCA密码子更具特异性，而UCA阅读效率没有显著下降。
