PROPERTIES OF A U1/MESSENGER-RNA 5' SPLICE SITE DUPLEX CONTAINING PSEUDOURIDINE AS MEASURED BY THERMODYNAMIC AND NMR METHODS
PROPERTIES OF A U1/MESSENGER-RNA 5' SPLICE SITE DUPLEX CONTAINING PSEUDOURIDINE AS MEASURED BY THERMODYNAMIC AND NMR METHODS
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DOI：
10.1021/bi00221a010
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发表时间：
1991-02-19
期刊：
影响因子：
2.9
通讯作者：
MCLAUGHLIN, LW
中科院分区：
文献类型：
--
作者：
HALL, KB;MCLAUGHLIN, LW
三种RNA十一聚体，5'AUAC-PSI-PSI-ACCUG （PSI =假尿嘧啶）。在控孔玻璃（CPG）载体上，采用亚磷酸酯三酯化学方法合成了5’auacuuaccug及其互补聚合物5’cagguaaguau。由这些分子形成的两个双链，5' auac - psi - psi - accug /5' cagguaaguau和5‘ auacuuaccug /5‘ cagguaaguau，代表了与mRNA一致的5’剪接位点配对的人类U1 snRNA的5’端。在一种脱氨基化合物中，假尿嘧啶被掺入到与体内原生U1 snRNA相对应的位置，而另一种（对照）脱氨基化合物仅含有尿嘧啶。令人惊讶的是，核磁共振数据显示，在主槽中发现的假尿嘧啶的额外亚质子可能不是氢键，在pH为6时的亚质子核磁共振谱中清晰可见。这一结果表明，RNA的结构限制了溶剂进入主槽，减缓了假尿嘧啶NH1亚氨基质子的交换。对两种双相化合物热力学性质的比较表明，以假尿嘧啶取代尿嘧啶后，双相化合物形成的自由能没有变化。
Three RNA undecamers, 5'AUAC-PSI-PSI-ACCUG (PSI = pseudouridine). 5'AUACUUACCUG, and their complementary ll-mer 5'CAGGUAAGUAU, have been chemically synthesized by phosphite triester chemistry on a controlled-pore glass (CPG) support. The two duplexes formed with these molecules, 5'AUAC-PSI-PSI-ACCUG/5'CAGGUAAGUAU and 5'AUACUUACCUG/5'CAGGUAAGUAU, represent the 5' end of human U1 snRNA paired to the mRNA consensus 5' splice site. In one undecamer, pseudouridines are incorporated at those positions corresponding to the native in vivo U1 snRNA, while the other (control) undecamer contains only uridine. Surprisingly, the NMR data show that the extra imino proton of the pseudouridines, which is found in the major groove and is presumably not hydrogen bonded, is clearly visible in the imino proton NMR spectrum at pH 6. This result suggests that the structure of the RNA restricts access of solvent to the major groove, slowing the exchange of the pseudouridine NH1 imino proton. A comparison of the thermodynamic properties of the two duplexes show that the free energy of duplex formation is unchanged by the substitution of pseudouridine for uridine.