Reconstitution of a functional IS608 single-strand transpososome: role of non-canonical base pairing.
Reconstitution of a functional IS608 single-strand transpososome: role of non-canonical base pairing.
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DOI：
10.1093/nar/gkr566
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发表时间：
2011-10
影响因子：
14.9
通讯作者：
Ton-Hoang B
中科院分区：
文献类型：
--
作者：
He S;Hickman AB;Dyda F;Johnson NP;Chandler M;Ton-Hoang B
单链（ss）转座是一种最近发现的由广泛分布的插入序列（IS）的IS 200/IS 605家族成员采用的机制，由转座酶TnpA催化。IS 608的转座酶识别IS两端的亚末端不完全回文序列（IP），并在位于一定距离处的位点处切割。切割位点C不直接被蛋白质识别，而是通过每个IP引导（G）序列的5′端的短序列识别，使用典型的（“沃森-克里克”）碱基相互作用网络。此外，还涉及一组类似于RNA结构中发现的非规范碱基相互作用。我们已经重建了一个生物相关的复合物，转座体，包括左右两端和TnpA，它催化切除的SS DNA环中间。我们提供了一个详细的图片的方式，其中IS 608转座体组装和证明，C和G序列是必不可少的，形成一个强大的转座体可检测的EMSA。我们还解决了几个问题的中心的SS转座体的组织和功能，并证明了非典型的碱基相互作用的IS 608结束其稳定性的重要作用，通过使用点突变破坏个人的非典型的碱基相互作用。
Single-stranded (ss) transposition, a recently identified mechanism adopted by members of the widespread IS200/IS605 family of insertion sequences (IS), is catalysed by the transposase, TnpA. The transposase of IS608, recognizes subterminal imperfect palindromes (IP) at both IS ends and cleaves at sites located at some distance. The cleavage sites, C, are not recognized directly by the protein but by short sequences 5′ to the foot of each IP, guide (G) sequences, using a network of canonical (‘Watson–Crick’) base interactions. In addition a set of non-canonical base interactions similar to those found in RNA structures are also involved. We have reconstituted a biologically relevant complex, the transpososome, including both left and right ends and TnpA, which catalyses excision of a ss DNA circle intermediate. We provide a detailed picture of the way in which the IS608 transpososome is assembled and demonstrate that both C and G sequences are essential for forming a robust transpososome detectable by EMSA. We also address several questions central to the organization and function of the ss transpososome and demonstrate the essential role of non-canonical base interactions in the IS608 ends for its stability by using point mutations which destroy individual non-canonical base interactions.
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影响因子：
14.9
作者：
Leontis, NB;Stombaugh, J;Westhof, E
通讯作者：
Westhof, E
影响因子：
16
作者：
Hickman, AB;Ronning, DR;Dyda, F
通讯作者：
Dyda, F
影响因子：
11.4
作者：
Bouvier, M;Demarre, G;Mazel, D
通讯作者：
Mazel, D
影响因子：
16
作者：
Guynet, Catherine;Achard, Adeline;Chandler, Michael
通讯作者：
Chandler, Michael
影响因子：
64.5
作者：
Barabas, Orsolya;Ronning, Donald R.;Dyda, Fred
通讯作者：
Dyda, Fred