Characterization of the trans Watson-Crick GU base pair located in the catalytic core of the antigenomic HDV ribozyme.
Characterization of the trans Watson-Crick GU base pair located in the catalytic core of the antigenomic HDV ribozyme.
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DOI：
10.1371/journal.pone.0040309
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Perreault JP
中科院分区：
文献类型：
--
作者：
Lévesque D;Reymond C;Perreault JP
HDV核酶的折叠途径是迄今为止针对小核酶阐明的最复杂的折叠途径。它包括6个不同的步骤，已被证明发生在化学裂解之前。很可能还有其他步骤有待发现。这些未知步骤中最关键的步骤之一是在环III内形成反式沃森-克里克GU碱基对。形成该碱基对的U23和G28核苷酸在HDV核酶的所有天然变体中完全保守，因此被认为是HDV样核酶的特征的一部分。该碱基对的形成和转化主要通过晶体结构和分子动力学模拟来研究。为了获得在溶液中形成该碱基对的物理支持，进行了一组实验，包括直接诱变、化学基团的位点特异性取代、动力学研究、化学探测和镁诱导的切割，其具体目标是表征反基因组HDV核酶中的该反式Watson-Crick GU碱基对。U23和G28都可以取代可能保留切割步骤之前和之后存在的一些H键相互作用的核苷酸。更稳定的反式沃森-克里克碱基对的形成被证明是一个后裂解事件，而一个可能较弱的反式沃森-克里克/Hoogsteen相互作用似乎在裂解步骤之前形成。这种异常稳定的切割后碱基对的形成可以通过有利于形成更稳定的产物-核酶复合物的基态而作为化学切割的驱动力。据我们所知，这是第一次证明了在核酶催化的反应中裂解后构象转换事件的潜在稳定作用。
The HDV ribozyme’s folding pathway is, by far, the most complex folding pathway elucidated to date for a small ribozyme. It includes 6 different steps that have been shown to occur before the chemical cleavage. It is likely that other steps remain to be discovered. One of the most critical of these unknown steps is the formation of the trans Watson-Crick GU base pair within loop III. The U23 and G28 nucleotides that form this base pair are perfectly conserved in all natural variants of the HDV ribozyme, and therefore are considered as being part of the signature of HDV-like ribozymes. Both the formation and the transformation of this base pair have been studied mainly by crystal structure and by molecular dynamic simulations. In order to obtain physical support for the formation of this base pair in solution, a set of experiments, including direct mutagenesis, the site-specific substitution of chemical groups, kinetic studies, chemical probing and magnesium-induced cleavage, were performed with the specific goal of characterizing this trans Watson-Crick GU base pair in an antigenomic HDV ribozyme. Both U23 and G28 can be substituted for nucleotides that likely preserve some of the H-bond interactions present before and after the cleavage step. The formation of the more stable trans Watson-Crick base pair is shown to be a post-cleavage event, while a possibly weaker trans Watson-Crick/Hoogsteen interaction seems to form before the cleavage step. The formation of this unusually stable post-cleavage base pair may act as a driving force on the chemical cleavage by favouring the formation of a more stable ground state of the product-ribozyme complex. To our knowledge, this represents the first demonstration of a potential stabilising role of a post-cleavage conformational switch event in a ribozyme-catalyzed reaction.
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影响因子：
14.9
作者：
Nehdi A;Perreault JP
通讯作者：
Perreault JP
DOI：
10.1089/oli.1.2000.10.53
发表时间：
2000-02-01
期刊：
ANTISENSE & NUCLEIC ACID DRUG DEVELOPMENT
影响因子：
--
作者：
Deschênes, P;Lafontaine, DA;Perreault, JP
通讯作者：
Perreault, JP
影响因子：
5.6
作者：
Krasovska, MV;Sefcikova, J;Walter, NG
通讯作者：
Walter, NG
影响因子：
14.9
作者：
Leontis, NB;Stombaugh, J;Westhof, E
通讯作者：
Westhof, E
影响因子：
64.8
作者：
Parisien, Marc;Major, Francois
通讯作者：
Major, Francois