Fan Chenguang, Ho Joanne M L, Chirathivat Napon, Söll Dieter, Wang Yane-Shih
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520 (USA).
Department of Chemistry, Yale University, New Haven, CT 06520 (USA).
Chembiochem. 2014 Aug 18;15(12):1805-1809. doi: 10.1002/cbic.201402083. Epub 2014 May 30.
We tested the substrate range of four wild-type E. coli aminoacyl-tRNA synthetases (AARSs) with a library of nonstandard amino acids (nsAAs). Although these AARSs could discriminate efficiently against the other canonical amino acids, they were able to use many nsAAs as substrates. Our results also show that E. coli tryptophanyl-tRNA synthetase (TrpRS) and tyrosyl-tRNA synthetase have overlapping substrate ranges. In addition, we found that the nature of the anticodon sequence of tRNA(Trp) altered the nsAA substrate range of TrpRS; this implies that the sequence of the anticodon affects the TrpRS amino acid binding pocket. These results highlight again that inherent AARS polyspecificity will be a major challenge in the aim of incorporating multiple different amino acids site-specifically into proteins.
我们用一个非标准氨基酸（nsAA）文库测试了四种野生型大肠杆菌氨酰-tRNA合成酶（AARS）的底物范围。尽管这些AARS能够有效地区分其他标准氨基酸，但它们能够使用许多nsAA作为底物。我们的结果还表明，大肠杆菌色氨酰-tRNA合成酶（TrpRS）和酪氨酰-tRNA合成酶具有重叠的底物范围。此外，我们发现tRNA(Trp)反密码子序列的性质改变了TrpRS的nsAA底物范围；这意味着反密码子序列会影响TrpRS的氨基酸结合口袋。这些结果再次突出表明，在将多种不同氨基酸位点特异性地掺入蛋白质的目标中，AARS固有的多特异性将是一个重大挑战。
