Maag David, Fekete Christie A, Gryczynski Zygmunt, Lorsch Jon R
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Mol Cell. 2005 Jan 21;17(2):265-75. doi: 10.1016/j.molcel.2004.11.051.
During eukaryotic translation initiation, ribosomal 43S complexes scan mRNAs for the correct AUG codon at which to begin translation. Start codon recognition triggers GTP hydrolysis, committing the complex to engagement at that point on the mRNA. While fidelity at this step is essential, the nature of the codon recognition event and the mechanism by which it activates GTP hydrolysis are poorly understood. Here we report the changes that occur within the 43S.mRNA complex in response to AUG codon recognition. eIF1 and eIF1A are key players in assembly of 43S.mRNA complexes capable of locating initiation codons. We observed FRET between these two factors when bound to the 40S subunit. Using steady-state FRET, anisotropy, and kinetic analyses, we demonstrate that start codon recognition results in a conformational change and release of eIF1 from the ribosome. These rearrangements probably play a role in triggering GTP hydrolysis and committing the complex to downstream events.
在真核生物翻译起始过程中，核糖体43S复合物扫描mRNA，寻找用于起始翻译的正确AUG密码子。起始密码子识别触发GTP水解，使复合物在mRNA上的该位点参与翻译。虽然这一步骤的保真度至关重要，但密码子识别事件的本质及其激活GTP水解的机制却知之甚少。在这里，我们报告了43S.mRNA复合物中因AUG密码子识别而发生的变化。eIF1和eIF1A是能够定位起始密码子的43S.mRNA复合物组装中的关键因子。当它们与40S亚基结合时，我们观察到这两个因子之间的荧光共振能量转移（FRET）。通过稳态FRET、各向异性和动力学分析，我们证明起始密码子识别导致构象变化以及eIF1从核糖体上释放。这些重排可能在触发GTP水解以及使复合物进入下游事件中发挥作用。
