Christian E L, Yarus M
Department of Molecular Cellular and Developmental Biology, University of Colorado, Boulder 80309-0347.
J Mol Biol. 1992 Dec 5;228(3):743-58. doi: 10.1016/0022-2836(92)90861-d.
We have developed a quantitative substitution interference technique to examine the role of Pro-Rp oxygens in the phosphodiester backbone of RNA, using phosphorothioates as a structural probe. This approach is generally applicable to any reaction involving RNA in which the precursor and reaction products can be separated. We have applied the technique to identity structural requirements in the group I intron from Tetrahymena thermophila for catalysis of hydrolysis at the 3' splice site; 44 phosphate oxygens are important in 3' splice site hydrolysis. These include four or five oxygens previously observed to be important in exon ligation. Although phosphate oxygens having a functional significance can be found throughout the intron, the strongest phosphorothioate effects are closely associated with positions in the highly conserved intron core, which are likely to be involved in tertiary interactions, substrate recognition and catalysis.
我们开发了一种定量取代干扰技术，以硫代磷酸酯作为结构探针，研究RNA磷酸二酯主链中Pro-Rp氧的作用。这种方法通常适用于任何涉及RNA的反应，前提是前体和反应产物能够分离。我们已将该技术应用于确定嗜热四膜虫I组内含子中3'剪接位点催化水解的结构要求；44个磷酸氧在3'剪接位点水解中起重要作用。其中包括先前观察到在 exon 连接中起重要作用的四个或五个氧。虽然在整个内含子中都能找到具有功能意义的磷酸氧，但最强的硫代磷酸酯效应与高度保守的内含子核心中的位置密切相关，这些位置可能参与三级相互作用、底物识别和催化。
