Escherichia coli tRNA(4)(Arg)(UCU) induces a constrained conformation of the crucial Omega-loop of arginyl-tRNA synthetase

Previous investigations show that tRNA(Arg)-induced conformational changes of arginyl-tRNA synthetase (ArgRS) Omega-loop region (Escherichia coli (E. coli), Ala451-Ala457) may contribute to the productive conformation of the enzyme catalytic core, and E. coli tRNA(2)(Arg)(ICG)-bound and -free conformations of the Omega-loop exchange at an intermediate rate on NMR timescale. Herein, we report that E. coli ArgRS catalyzes tRNA(2)(Arg)(ICG) and tRNA(4)(Arg)(UCU) with similar efficiencies. However, 19F NMR spectroscopy of 4-fluorotryptophan-labeled E. coli ArgRS reveals that the tRNA(4)(Arg)(UCU)-bound and -free conformations of the Omega-loop region interconvert very slowly and the lifetime of bound conformation is much longer than 0.33 ms. Therefore, tRNA(4)(Arg)(UCU) differs from tRNA(2)(Arg)(ICG) in the conformation-exchanging rate of the Omega-loop. Comparative structure model of E. coli ArgRS is presented to rationalize these 19F NMR data. Our 19F NMR and catalytic assay results suggest that the tRNA(Arg)-induced conformational changes of Omega-loop little contribute to the productive conformation of ArgRS catalytic core.     

稀有密码子对proUK基因在大肠杆菌中高表达的影响

ＡＧＧ／ＡＧＡ密码子在ｐｒｏＵＫ基因中出现的频率高达２％,大肠杆菌中现有的编码ｔＲＮＡＵＣＵ的ａｒｇＵ基因不能满足ｐｒｏＵＫ高表达的需要。本文研究了ａｒｇＵ基因剂量对ｐｒｏＵＫ表达的影响,结果表明带有ａｒｇＵ基因的中等拷贝数质粒能促进ｐｒｏＵＫ基因表达。