RNA cleavage by a DNA enzyme with extended chemical functionality
RNA cleavage by a DNA enzyme with extended chemical functionality
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DOI：
10.1021/ja993688s
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发表时间：
2000-03-22
影响因子：
15
通讯作者：
Barbas, CF
中科院分区：
文献类型：
--
作者：
Santoro, SW;Joyce, GF;Barbas, CF
在体外选择技术被应用到开发的DNA酶，含有三个催化必需的咪唑基团和催化裂解的RNA底物。通过聚合酶催化掺入C5-咪唑官能化脱氧尿苷代替胸苷构建用于选择的核酸文库。化学合成用于定义仅由12个残基组成的最小化催化结构域。催化结构域形成一个紧凑的发夹结构，显示三个含咪唑的残基。通过简单改变催化结构域周围的两个底物识别结构域，该酶可以切割几乎任何序列的RNA。该酶在微摩尔浓度的Zn 2+存在下以多个周转操作，表现出饱和动力学和> 1 min（-1）的催化速率。含咪唑的DNA酶是已知的最小的核酸酶之一，它将核酸酶的底物识别特性和蛋白酶的化学功能性结合在一个小分子中，但它是通用的和催化有效的。
In vitro selection techniques were applied to the development of a DNA enzyme that contains three catalytically essential imidazole groups and catalyzes the cleavage of RNA substrates. Nucleic acid libraries for selection were constructed by polymerase-catalyzed incorporation of C5-imidazole-functionalized deoxyuridine in place of thymidine. Chemical synthesis was used to define a minimized catalytic domain composed of only 12 residues. The catalytic domain forms a compact hairpin structure that displays the three imidazole-containing residues. The enzyme can be made to cleave RNAs of almost any sequence by simple alteration of the two substrate-recognition domains that surround the catalytic domain. The enzyme operates with multiple turnover in the presence of micromolar concentrations of Zn2+, exhibiting saturation kinetics and a catalytic rate of > 1 min(-1). The imidazole-containing DNA enzyme, one of the smallest known nucleic acid enzymes, combines the substrate-recognition properties of nucleic acid enzymes and the chemical functionality of protein enzymes in a molecule that is small, yet versatile and catalytically efficient.