通常不受欢迎的 RNA 水解反应的 DNA 催化,Journal of the American Chemical Society

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通常不受欢迎的 RNA 水解反应的 DNA 催化
Journal of the American Chemical Society ( IF 14.4 ) Pub Date : 2013-06-03 , DOI: 10.1021/ja4032488
Darren J Parker ₁ , Ying Xiao , John M Aguilar , Scott K Silverman

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我们最近使用体外选择来鉴定许多催化 DNA 磷酸二酯键水解并产生 5'-磷酸和 3'-羟基末端的脱氧核酶。或者，已鉴定出许多脱氧核酶可通过 2'-羟基酯交换催化 RNA 裂解，形成 2',3'-环磷酸酯和 5'-羟基末端。在这项研究中，我们研究了 DNA 通过水解而不是酯交换来催化 RNA 切割的能力，尽管通常水解反应相对于酯交换来说是非常不利的。通过一系列体外选择实验，我们发现重新选择 DNA 水解脱氧核酶会导致酯交换或水解，这取决于排除或包含严格的水解选择压力。从随机 DNA 库开始的全新选择，使用全 RNA 底物并施加相同的选择压力，也导致 RNA 水解。总的来说，这些结果通过实验证明小 DNA 序列具有将化学反应引导到不利途径的催化能力，即使更有利的机制很容易获得。因此，我们对 DNA 催化的看法扩展到不仅仅是提高没有催化剂的情况下会发生得更慢的反应速率。即使更有利的机制很容易获得。因此，我们对 DNA 催化的看法扩展到不仅仅是提高没有催化剂的情况下会发生得更慢的反应速率。即使更有利的机制很容易获得。因此，我们对 DNA 催化的看法扩展到不仅仅是增加没有催化剂的情况下会发生得更慢的反应速率。

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DNA Catalysis of a Normally Disfavored RNA Hydrolysis Reaction

We recently used in vitro selection to identify many deoxyribozymes that catalyze DNA phosphodiester bond hydrolysis and create 5'-phosphate and 3'-hydroxyl termini. Alternatively, numerous deoxyribozymes have been identified for catalysis of RNA cleavage by 2'-hydroxyl transesterification, forming 2',3'-cyclic phosphate and 5'-hydroxyl termini. In this study, we investigated the ability of DNA to catalyze RNA cleavage by hydrolysis rather than transesterification, although normally the hydrolysis reaction is substantially disfavored relative to transesterification. Via a series of in vitro selection experiments, we found that reselection of a DNA-hydrolyzing deoxyribozyme leads either to transesterification or hydrolysis, depending on exclusion or inclusion of a stringent selection pressure for hydrolysis. An entirely new selection starting from a random DNA pool, using an all-RNA substrate and imposing the same selection pressure, also leads to RNA hydrolysis. Collectively, these results establish experimentally that small DNA sequences have the catalytic ability to direct a chemical reaction down a disfavored pathway, even when a more favorable mechanism is readily available. Our view of DNA catalysis is therefore expanded beyond merely increasing the rates of reactions that would have occurred more slowly without the catalyst.

更新日期：2013-06-03

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