A general purpose RNA-cleaving DNA enzyme
A general purpose RNA-cleaving DNA enzyme
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DOI：
10.1073/pnas.94.9.4262
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发表时间：
1997-04-29
影响因子：
11.1
通讯作者：
Joyce, GF
中科院分区：
文献类型：
--
作者：
Santoro, SW;Joyce, GF
利用体外选择程序开发了一种DNA酶，该酶可以在模拟生理条件下切割几乎任何目标RNA底物，该酶由15个脱氧核苷酸的催化结构域组成，两侧是两个底物识别结构域，每个结构域含有7至8个脱氧核苷酸。RNA底物通过沃森-克里克碱基配对，并在位于未配对嘌呤和配对嘧啶残基之间的特定磷酸二酯处被切割。尽管DNA酶体积小，但在多种周转条件下，其催化效率（k(cat)/ k -m）接近10(9)M-1 min(-1)，超过任何其他已知的核酸酶，其活性依赖于Mg2+离子的存在。通过改变底物识别结构域的序列，DNA酶可以靶向不同的RNA底物。例如，在本研究中，它被用于切割HIV-1 gag/pol、env、vpr、that和nef mrna起始密码子区域对应的合成rna。
An in vitro selection procedure was used to develop a DNA enzyme that can be made to cleave almost any targeted RNA substrate under simulated physiological conditions, The enzyme is comprised of a catalytic domain of 15 deoxynucleotides, flanked by two substrate-recognition domains of seven to eight deoxynucleotides each. The RNA substrate is hound through Watson-Crick base pairing and is cleaved at a particular phosphodiester located between an unpaired purine and a paired pyrimidine residue, Despite its small size, the DNA enzyme has a catalytic efficiency (k(cat)/K-m) of approximate to 10(9) M-1 min(-1) under multiple turnover conditions, exceeding that of any other known nucleic acid enzyme, Its activity is dependent on the presence of Mg2+ ion. By changing the sequence of the substrate-recognition domains, the DNA enzyme cars be made to target different RNA substrates. In this study, for example, it was directed to cleave synthetic RNAs corresponding to the start codon region of HIV-1 gag/pol, env, vpr, tat, and nef mRNAs.