Dotson P Patrick, Johnson Ashley K, Testa Stephen M
Department of Chemistry, University of Kentucky, Lexington, KY 40506, USA.
Nucleic Acids Res. 2008 Sep;36(16):5281-9. doi: 10.1093/nar/gkn507. Epub 2008 Aug 6.
Group I intron-derived ribozymes can catalyze a variety of non-native reactions. For the trans-excision-splicing (TES) reaction, an intron-derived ribozyme from the opportunistic pathogen Pneumocystis carinii catalyzes the excision of a predefined region from within an RNA substrate with subsequent ligation of the flanking regions. To establish TES as a general ribozyme-mediated reaction, intron-derived ribozymes from Tetrahymena thermophila and Candida albicans, which are similar to but not the same as that from Pneumocystis, were investigated for their propensity to catalyze the TES reaction. We now report that the Tetrahymena and Candida ribozymes can catalyze the excision of a single nucleotide from within their ribozyme-specific substrates. Under the conditions studied, the Tetrahymena and Candida ribozymes, however, catalyze the TES reaction with lower yields and rates [Tetrahymena (k(obs)) = 0.14/min and Candida (k(obs)) = 0.34/min] than the Pneumocystis ribozyme (k(obs) = 3.2/min). The lower yields are likely partially due to the fact that the Tetrahymena and Candida catalyze additional reactions, separate from TES. The differences in rates are likely partially due to the individual ribozymes ability to effectively bind their 3' terminal guanosines as intramolecular nucleophiles. Nevertheless, our results demonstrate that group I intron-derived ribozymes are inherently able to catalyze the TES reaction.
I 组内含子衍生的核酶可以催化多种非天然反应。对于反式切除-剪接(TES)反应,来自机会性病原体卡氏肺孢子虫的一种内含子衍生核酶催化从RNA底物内部切除一个预定义区域,随后将侧翼区域连接起来。为了确定TES作为一种普遍的核酶介导反应,研究了来自嗜热四膜虫和白色念珠菌的内含子衍生核酶,它们与卡氏肺孢子虫的核酶相似但不同,考察它们催化TES反应的倾向。我们现在报告,四膜虫和念珠菌核酶可以催化从其核酶特异性底物内部切除单个核苷酸。然而,在所研究的条件下,四膜虫和念珠菌核酶催化TES反应的产率和速率 [四膜虫(k(obs))= 0.14/分钟,念珠菌(k(obs))= 0.34/分钟] 低于卡氏肺孢子虫核酶(k(obs) = 3.2/分钟)。产率较低可能部分是由于四膜虫和念珠菌催化了与TES不同的其他反应。速率差异可能部分是由于各个核酶有效结合其3'末端鸟苷作为分子内亲核试剂的能力。尽管如此,我们的结果表明,I组内含子衍生的核酶本质上能够催化TES反应。