High-Throughput Mutational Analysis of a Twister Ribozyme.
High-Throughput Mutational Analysis of a Twister Ribozyme.
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DOI：
10.1002/anie.201605470
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发表时间：
2016-08-22
影响因子：
16.6
通讯作者：
Yokobayashi, Yohei
中科院分区：
文献类型：
--
作者：
Kobori, Shungo;Yokobayashi, Yohei
最近在不同生物体中发现了新类型的自切割核酶，引发了人们对核酶化学和生物学的新兴趣。核酶的功能分析和工程化通常涉及对多个核酶突变体进行生化测定。然而，由于每个核酶突变体必须单独制备和测定，可以研究的突变体的数量和种类受到严重限制。产生扭转酶核酶的所有单突变体和双突变体（总共10296个突变体），并通过利用深度测序来计数每个突变体的切割和未切割序列的数量来测定它们的自切割活性。有趣的是，我们发现核酶对突变是高度稳健的，使得在测定条件下分别有71%和30%的所有单突变体和双突变体保留可检测的活性。还观察到，构成核酶的结构元件对突变表现出不同的敏感性。
Recent discoveries of new classes of self‐cleaving ribozymes in diverse organisms have triggered renewed interest in the chemistry and biology of ribozymes. Functional analysis and engineering of ribozymes often involve performing biochemical assays on multiple ribozyme mutants. However, because each ribozyme mutant must be individually prepared and assayed, the number and variety of mutants that can be studied are severely limited. All of the single and double mutants of a twister ribozyme (a total of 10 296 mutants) were generated and assayed for their self‐cleaving activity by exploiting deep sequencing to count the numbers of cleaved and uncleaved sequences for every mutant. Interestingly, we found that the ribozyme is highly robust against mutations such that 71 % and 30 % of all single and double mutants, respectively, retain detectable activity under the assay conditions. It was also observed that the structural elements that comprise the ribozyme exhibit distinct sensitivity to mutations.
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影响因子：
14.8
作者：
Weinberg Z;Kim PB;Chen TH;Li S;Harris KA;Lünse CE;Breaker RR
通讯作者：
Breaker RR
影响因子：
14.8
作者：
Roth, Adam;Weinberg, Zasha;Chen, Andy G. Y.;Kim, Peter B.;Ames, Tyler D.;Breaker, Ronald R.
通讯作者：
Breaker, Ronald R.
影响因子：
15
作者：
Wilson, Timothy J.;Liu, Yijin;Liiley, David M. J.
通讯作者：
Liiley, David M. J.
DOI：
10.1073/pnas.1414571111
发表时间：
2014-09-09
影响因子：
11.1
作者：
Eiler, Daniel;Wang, Jimin;Steitz, Thomas A.
通讯作者：
Steitz, Thomas A.
影响因子：
2.9
作者：
Ucisik, Melek N.;Bevilacqua, Philip C.;Hammes-Schiffer, Sharon
通讯作者：
Hammes-Schiffer, Sharon