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首页> 外文期刊>RSC Advances >RNA ligation of very small pseudo nick structures by T4 RNA ligase 2, leading to efficient production of versatile RNA rings
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RNA ligation of very small pseudo nick structures by T4 RNA ligase 2, leading to efficient production of versatile RNA rings

机译：T4 RNA连接酶2可以将非常小的假缺口结构进行RNA连接，从而可以有效生产通用的RNA环

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摘要

T4 RNA ligase 2 catalyses two types of reactions: (i) sealing of a nick structure in double-stranded RNA and (ii) connection of two single-stranded RNA strands. In order to obtain comprehensive views on these two types of reactions and widen the application scope of this RNA ligase, we here systematically analysed the connection of single-stranded RNA strands having different secondary structures. It has been found that the ligation is enormously promoted when a stem of only 4-bp or longer is formed in the 3′-OH side of the joining site. Additional placement of a stem in the 5′-phosphate side further facilitates the ligation. In contrast, perturbation of the stem structures in RNA substrates suppresses the ligation. These results indicate that ligation of two single-stranded RNA strands by T4 RNA ligase 2 is greatly promoted by forming a “nick-like intermediate”. Even the unstable intermediate, formed only temporarily in the solution, is sufficiently effective. By designing the synthetic systems in terms of this finding, short single-stranded RNA rings of versatile sizes, which are otherwise hard to be obtained, are efficiently prepared in high selectivity and yield.
机译：T4 RNA连接酶2催化两种类型的反应：（i）双链RNA中的缺口结构密封，以及（ii）两根单链RNA链的连接。为了获得对这两种反应的全面了解并扩大该RNA连接酶的应用范围，我们在这里系统地分析了具有不同二级结构的单链RNA链的连接。已经发现，当在连接位点的3'-OH侧仅形成4-bp或更长的茎时，极大地促进了连接。将茎在5'-磷酸侧的额外放置进一步促进了连接。相反，RNA底物中茎结构的扰动会抑制连接。这些结果表明，通过形成“缺口状中间体”，大大促进了T4 RNA连接酶2对两条单链RNA链的连接。即使是仅在溶液中临时形成的不稳定中间体也足够有效。通过根据这一发现设计合成系统，可以以高选择性和高收率有效地制备原本难以获得的通用尺寸的短单链RNA环。

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