Mechanism for the definition of elongation and termination by the class II CCA-adding enzyme
Mechanism for the definition of elongation and termination by the class II CCA-adding enzyme
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DOI：
10.1038/emboj.2009.260
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发表时间：
2009-11-04
期刊：
影响因子：
11.4
通讯作者：
Tomita, Kozo
中科院分区：
文献类型：
--
作者：
Toh, Yukimatsu;Takeshita, Daijiro;Tomita, Kozo
CCA添加酶在没有核酸模板的情况下在tRNA的3'末端合成CCA序列。测定了II类海栖热袍菌（Thermotoga maritima）CCA加成酶及其与CTP或ATP形成的复合物的晶体结构。基于结构的替代的催化头和核碱基相互作用的颈部结构域的遗传学密切相关的Aquifex aeolicus A-添加酶的相应结构域的T。海产CCA加酶使A加酶在体内和体外都能加CCA。然而，仅催化头部结构域的替换不允许A-添加酶添加CCA，并且该酶表现出（A，C）-添加活性。我们确定了颈部结构域中阻止（A，C）添加活性的区域，并定义了核苷酸重复的数量和正确CCA添加的特异性。我们还确定了头部结构域中的区域，该区域定义了CC添加后的末端A添加。这些结果共同表明，在II类CCA添加酶中，头部和颈部结构域协同地和动态地定义核苷酸添加的数量和核苷酸选择的特异性。The EMBO Journal（2009）28，3353-3365. doi：10.1038/doj.2009.260; 2009年9月10日在线发布
The CCA-adding enzyme synthesizes the CCA sequence at the 3' end of tRNA without a nucleic acid template. The crystal structures of class II Thermotoga maritima CCA-adding enzyme and its complexes with CTP or ATP were determined. The structure-based replacement of both the catalytic heads and nucleobase-interacting neck domains of the phylogenetically closely related Aquifex aeolicus A-adding enzyme by the corresponding domains of the T. maritima CCA-adding enzyme allowed the A-adding enzyme to add CCA in vivo and in vitro. However, the replacement of only the catalytic head domain did not allow the A-adding enzyme to add CCA, and the enzyme exhibited (A, C)-adding activity. We identified the region in the neck domain that prevents (A, C)-adding activity and defines the number of nucleotide incorporations and the specificity for correct CCA addition. We also identified the region in the head domain that defines the terminal A addition after CC addition. The results collectively suggest that, in the class II CCA-adding enzyme, the head and neck domains collaboratively and dynamically define the number of nucleotide additions and the specificity of nucleotide selection. The EMBO Journal (2009) 28, 3353-3365. doi: 10.1038/emboj.2009.260; Published online 10 September 2009