The molecular mechanism of post-peptidyl quality control on the ribosome

项目概要： 生物体的生长和生存能力依赖于其基因组的忠实和快速解码 信息转化为功能性肽序列。高精度的蛋白质合成确保了错误的 多肽，其更倾向于错误折叠并因此可能具有不希望的毒性后果， 没有生产。蛋白质合成的整体保真度似乎受到蛋白质合成的作用的限制。 核糖体，这是两个亚基大分子机器负责解码的 信使RNA转化为蛋白质在生命的各个领域。在每个延伸周期中， 仔细选择与解码中的密码子匹配的适当氨酰-tRNA（aa-tRNA）， 从大量相互竞争的aa-tRNA中分离出来。除此之外，我们最近还发现了一个 在肽键形成后发生的核糖体质量控制机制， 有助于高保真蛋白质合成。类似于DNA所享有的校对策略， RNA聚合酶和tRNA合成酶，新发现的基于核糖体的机制已经就位 以监控刚刚完成的化学步骤的质量。在伸长周期期间，掺入 一个不正确的氨基酸被发现对随后的特异性有显着的影响， 反应这种错误的反复积累导致蛋白质合成的失败终止， 释放因子，其在正常条件下很少解码有义密码子。我们的长期目标 我们的工作是彻底了解这一过程的分子机制。我们 当前的目标是找出信号是如何从干扰的mRNA-tRNA相互作用中传递的 到解码中心，最终导致低保真度的蛋白质合成。我们也有兴趣 释放因子的活性如何在干扰的mRNA存在下在有义密码子上调节， tRNA相互作用，以及负责这种活动的结构线索。这些目标是建立在 在突变翻译组分的背景下，围绕前稳态动力学方法，以及 低分辨率结构探测技术。作为第三个目标，我们有兴趣探索一个以前 释放因子3在质量控制机制中的作用及其在细胞活力中的效用未知。 最后，我们感兴趣的是发现这个系统是否存在于真核生物中，并确定其他 在这个过程中可能涉及的因素，如果有的话。

Project Summary: Organismal growth and viability is dependent on the faithful and fast decoding of its genomic information into functional peptide sequences. High-accuracy protein synthesis ensures that errant polypeptides, which are more prone to misfold and hence may have undesirable toxic consequences, are not produced. The overall fidelity of protein synthesis appears to be limited by the action of the ribosome, which is the two-subunit macromolecular machine responsible for the decoding of the messenger RNA into protein in all domains of life. During each cycle of elongation, the ribosome carefully selects the appropriate aminoacyl-tRNA (aa-tRNA) that matches the codon in the decoding center from a large-pool of competing aa-tRNAs. In addition to this, we have recently uncovered a quality control mechanism on the ribosome that takes place after peptide-bond formation, which contributes to high-fidelity protein synthesis. Akin to the proofreading strategies enjoyed by DNA and RNA polymerases and tRNA synthetases, the newly discovered ribosome-based mechanism is in place to monitor the quality of the just completed chemical step. During the elongation cycle, the incorporation of an incorrect amino acid was found to have dramatic effects on the specificity of the subsequent reaction. This iterated accumulation of errors results in the abortive termination of protein synthesis by release factors, which under normal conditions rarely decode sense codons. The long term goal of our work is to gain a thorough understanding of the molecular mechanisms underlying this process. Our immediate goal is to find out how the signal is communicated from a perturbed mRNA-tRNA interaction to the decoding center, which ultimately leads to low-fidelity protein synthesis. We are also interested in how the activity of release factors is modulated on sense codons in the presence of a perturbed mRNA- tRNA interaction, and the structural cues that are responsible for this activity. These goals are built around pre-steady state kinetics approaches in the context of mutated translation components, and low-resolution structural probing techniques. As a third goal we are interested in exploring a previously unknown role for release factor 3 in the quality control mechanism and its utility in cellular viability. Finally we are interested in finding whether this system exists in eukaryotes, and identifying other factors, if any, that might be involved during this process.