Kinetics and thermodynamics of RRF, EF-G, and thiostrepton interaction on the Escherichia coli ribosome.
Kinetics and thermodynamics of RRF, EF-G, and thiostrepton interaction on the Escherichia coli ribosome.
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RRF、EF-G 和硫链丝菌素在大肠杆菌核糖体上相互作用的动力学和热力学。
DOI：
10.1021/bi048927p
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Cooperman,BarryS
中科院分区：
文献类型：
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作者：
Seo,Hyuk-Soo;Kiel,Michael;Pan,Dongli;Raj,VSamuel;Kaji,Akira;Cooperman,BarryS
核糖体再循环因子（RRF）和延伸因子-G（EF-G）共同对蛋白质合成终止后细菌核糖体的再循环至关重要。在这里，我们提出了平衡和快速的动力学测量允许制定一个最小的动力学计划，占定量的RRF和EF-G相互作用theEscherichia coliribosome。RRF和EF-G（a）在与裸核糖体结合时各自形成二元复合物，其经历异构化以形成更稳定的复合物，（B）在核糖体上形成混合三元复合物，其中对每种因子的亲和力显著低于其与裸核糖体结合的亲和力，和（c）各自与每个核糖体的两个位点结合，EF-G具有比RRF显著更高的第二位点亲和力。在核糖体-RRF复合物中加入EF-G可诱导RRF快速解离，解离速率与体内核糖体再循环速率一致，但加入RRF不会增加核糖体结合EF-G的不稳定性。添加的硫链丝菌素减慢EF-G的初始结合，并防止更稳定的EF-G复合物的形成和EF-G诱导的RRF解离。这些发现与终止后复杂拆卸的机制有关。
Ribosome recycling factor (RRF) and elongation factor-G (EF-G) are jointly essential for recycling bacterial ribosomes following termination of protein synthesis. Here we present equilibrium and rapid kinetic measurements permitting formulation of a minimal kinetic scheme that accounts quantitatively for RRF and EF-G interaction on theEscherichia coliribosome. RRF and EF-G (a) each form a binary complex on binding to a bare ribosome which undergoes isomerization to a more stable complex, (b) form mixed ternary complexes on the ribosome in which the affinity for each factor is considerably lower than its affinity for binding to a bare ribosome, and (c) each bind to two sites per ribosome, with EF-G having considerably higher second-site affinity than RRF. Addition of EF-G to the ribosome−RRF complex induces rapid RRF dissociation, at a rate compatible with the rate of ribosome recyclingin vivo, but added RRF does not increase the lability of ribosome-bound EF-G. Added thiostrepton slows the initial binding of EF-G, and prevents both formation of the more stable EF-G complex and EF-G-induced RRF dissociation. These findings are relevant for the mechanism of post-termination complex disassembly.