Protonation of non-Watson-Crick base pairs and encapsidation of turnip yellow mosaic virus RNA
Protonation of non-Watson-Crick base pairs and encapsidation of turnip yellow mosaic virus RNA
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DOI：
10.1073/pnas.202287499
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发表时间：
2002-10-15
影响因子：
11.1
通讯作者：
Pleij, CWA
中科院分区：
文献类型：
--
作者：
Bink, HHJ;Hellendoorn, K;Pleij, CWA
芜菁黄花叶病毒RNA的5'非编码区含有两个保守的发夹结构，其内环由CC和CA错配组成。在这篇文章中，有证据表明，5'近端发夹功能作为一个启动信号。对该发夹结构的广泛诱变研究和病毒后代的测序显示出对内环内C-C和C-A错配的明显偏好。这些错配的重要性在于它们的pH依赖性质子化和稳定的碱基对形成。发现在5'近端发夹的内环中缺乏可质子化错配的几种突变体的包封效率受到严重影响。此外，用各种RNA发夹和具有孔的空衣壳进行凝胶迁移率变化测定。含有（CC）-C-的可质子化发夹。和/或C（.）发现在亚精胺存在下，在酸性条件下（pH 4.5），A对特异性结合到蛋白质壳的内部。基于这些结果，我们提出，这种结合的质子化胞嘧啶的萝卜黄花叶病毒外壳蛋白可能代表一个新的主题在RNA-蛋白质相互作用。
The 5' UTR of turnip yellow mosaic virus RNA contains two conserved hairpins with internal loops consisting of CC and CA mismatches. In this article, evidence is presented indicating that the 5' proximal hairpin functions as an encapsidation initiation signal. Extensive mutagenesis studies on this hairpin and sequencing of virus progeny showed a clear preference for C-C and C-A mismatches within the internal loop. The importance of these mismatches lies in their pH-dependent protonation and stable base pair formation. Encapsidation efficiency was found to be severely affected for several mutants lacking the protonatable mismatches in the internal loop of the 5' proximal hairpin. Furthermore, gel mobility-shift assays were performed with various RNA hairpins and empty capsids with a hole. Protonatable hairpins containing (CC)-C-. and/or C(.)A pairs were found to bind specifically to the interior of the protein shell under acidic conditions (pH 4.5) in the presence of spermidine. Based on these results we propose that this binding of protonated cytosines to the coat protein of turnip yellow mosaic virus may represent a new motif in RNA-protein interactions.