5-METHYLCYTIDINE IS REQUIRED FOR COOPERATIVE BINDING OF MG2+ AND A CONFORMATIONAL TRANSITION AT THE ANTICODON STEM-LOOP OF YEAST PHENYLALANINE TRANSFER-RNA
5-METHYLCYTIDINE IS REQUIRED FOR COOPERATIVE BINDING OF MG2+ AND A CONFORMATIONAL TRANSITION AT THE ANTICODON STEM-LOOP OF YEAST PHENYLALANINE TRANSFER-RNA
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DOI：
10.1021/bi00089a047
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发表时间：
1993-09-28
期刊：
影响因子：
2.9
通讯作者：
AGRIS, PF
中科院分区：
文献类型：
--
作者：
CHEN, Y;SIERZPUTOWSKAGRACZ, H;AGRIS, PF
修饰核苷在tRNA结构和离子结合中的作用已经用对应于酵母tRNA（Phe）反密码子茎和环tRNA（AC）Phe）的化学合成RNA进行了研究。在tRNA（AC）Phe-d（m5 C-14），CCAGACUGAA-GAU-d（m5 C-14）-UGG的茎的位置14处引入d（m5 C），类似于天然tRNA（Phe）中的m5 C40，在远离m5 C的位点处引入强Mg 2+结合。通过圆二色性光谱检测到的Mg 2+诱导的结构转变类似于对tRNA（AC）Phe的DNA类似物观察到的结构转变（Guenther等人，1992; Dao等人，1992年）。相比之下，Mg 2+对未修饰的tRNA（AC）Phe-rC-14或tRNA（AC）Phe-d（C-14）几乎没有影响。 修饰的tRNA（AC）Phe-d（m5 C-14）结合两个Mg ~（2+）离子，且结合是协同的。tRNA（AC）Phe-d（m5 C-14）的两个Mg 2+离子的解离常数为2.5 × 10（-9）M2，这是由于Mg 2+结合显著稳定了RNA结构，DELTAG = -11.7 kcal/mol。通过H-1 NMR研究的tRNA（AC）Phe-d（m5 C-14）结构具有五个碱基对的双链茎和两个额外的碱基对，其跨越未修饰的tRNA（AC）Phe中的七元环。 酵母tRNA（AC）Phe中胞苷的甲基化使得该分子能够通过受Mg 2+浓度调节的过程形成一种以上的构象。因此，最简单的转录后修饰的tRNA，甲基化，是涉及一个有点远，内部网站镁2+结合和稳定的tRNA结构，特别是反密码子茎和环。
The role of modified nucleosides in tRNA structure and ion binding has been investigated with chemically synthesized RNAs corresponding to the yeast tRNA(Phe) anticodon stem and loop tRNA(AC)Phe). Incorporation of d(m5C) at position 14 of the stem of tRNA(AC)Phe-d(m5C-14), CCAGACUGAA-GAU-d(m5C-14)-UGG, analogous to m5C40 in native tRNA(Phe), introduced a strong Mg2+ binding at a site distant from the m5C. A Mg2+-induced structural transition, detected by circular dichroism spectroscopy, was similar to that observed for the DNA analog of tRNA(AC)Phe (Guenther et al., 1992; Dao et al., 1992). In contrast, Mg2+ had little effect on unmodified tRNA(AC)Phe-rC-14 or tRNA(AC)Phe-d(C-14). Modified tRNA(AC)Phe-d(m5C-14) bound two Mg2+ ions, and the binding was cooperative. The dissociation constant of the two Mg2+ ions from tRNA(AC)Phe-d(m5C-14), 2.5 X 10(-9) M2, is the result of an RNA structure significantly stabilized by Mg2+ binding, DELTAG = -11.7 kcal/mol. The tRNA(AC)Phe-d(m5C-14) structure, investigated by H-1 NMR, had a double stranded stem of five base pairs and two additional base pairs across what was a seven membered loop in the unmodified tRNA(AC)Phe. Methylation of cytidine in the yeast tRNA(AC)Phe enables the molecule to form more than one conformation through a process regulated by Mg2+ concentration. Thus, the simplest of posttranscriptional modifications of tRNA, a methylation, is involved in a somewhat distant, internal-site Mg2+ binding and stabilization of tRNA structure, especially that of the anticodon stem and loop.