The crystal structure and mutational analysis of a novel RNA-binding domain found in the human Tap nuclear mRNA export factor
The crystal structure and mutational analysis of a novel RNA-binding domain found in the human Tap nuclear mRNA export factor
复制标题
DOI：
10.1073/pnas.042698599
复制
发表时间：
2002-02-19
影响因子：
11.1
通讯作者：
Georgiadis, MM
中科院分区：
文献类型：
--
作者：
Ho, DN;Coburn, GA;Georgiadis, MM
Tap 蛋白介导细胞 mRNA 的序列非特异性核输出以及带有组成型转运元件 (CTE) 的逆转录病毒 mRNA 的序列特异性输出。之前，已经描述了各个 Tap 子结构域的结构，包括核糖核蛋白和富含亮氨酸的重复结构域。在这里，我们报道了人 Tap 的功能性 CTE RNA 结合结构域的晶体结构，包括核糖核蛋白结构域的 N 端臂和结构域间连接多肽。为了识别与 CTE 相互作用的残基，我们在 Tap CTE 结合域中引入了 38 个丙氨酸取代表面残基，并测试了这些突变体支持 CTE 依赖性核 RNA 输出和 CTE 结合的能力。发现在富含亮氨酸的重复结构域中的凹表面上聚集的四个残基对于 CTE 结合至关重要，并定义了该结构域上的 CTE 相互作用表面。 Tap 上第二个关键的 CTE 相互作用表面由核糖核蛋白结构域表面上三个先前识别的残基定义。结构和突变数据定义了 Tap 蛋白上的一个新的 RNA 结合位点。
The Tap protein mediates the sequence nonspecific nuclear export of cellular mRNAs as well as the sequence-specific export of retroviral mRNAs bearing the constitutive transport element (CTE). Previously, the structures of individual Tap subdomains, including ribonucleoprotein and leucine-rich repeat domains, have been described. Here, we report the crystal structure of a functional CTE RNA-binding domain of human Tap, including the N-terminal arm of the ribonucleoprotein domain and interdomain linking polypeptide. To identify residues that interact with the CTE, we have introduced 38 alanine substitutions for surface residues in the Tap CTE-binding domain and tested these mutants for their ability to support CTE-dependent nuclear RNA export and CTE binding. Four residues that cluster on a concave surface in the leucine-rich repeat domain were found to be critical for CTE binding and define a CTE-interacting surface on this domain. The second critical CTE-interacting surface on Tap is defined by three previously identified residues on the surface of the ribonucleoprotein domain. The structural and mutational data define a novel RNA-binding site on the Tap protein.