Preparation of Artificial nuclease and creation of new-biotechnology

DNA的非酶水解一直是化学家们研究的重要目标之一。早在几年前，人们就发现了稀土离子的显著催化作用，在pH为7的条件下，DNA中高度稳定的磷酸二酯键被水解，其中Ce(IV)离子的催化活性尤为显著。然而，Ce(IV)很容易形成金属氢氧化物凝胶，这一特点限制了其应用范围。具有催化活性的均相Ce(IV)络合物具有更广泛的应用前景。在这项研究中，我们研究了(1)催化活性的Ce(IV)配合物和(2)激活核酸中目标磷酸二酯的方法：(1)EDTA的均相Ce(IV)配合物在生理条件下能迅速水解寡核苷酸。相反，二核苷酸的水解度不会达到可测量的程度。DNA底物的聚合度对目前的催化是至关重要的。该络合物的活性通过与所选择的寡胺的协同作用而显著提高。在有精胺(10μ)存在时，反应速度约为无精胺时的50倍。(2)非共价体系(三元体系)，由(1)含吖啶的脱氧核糖核酸低聚物，(2)未修饰的脱氧核糖核酸，(3)游离稀土(III)离子组成的非共价体系(三元体系)，选择性地有效地将核糖核酸水解到目标双酯上。相应的磷酸二酯键被与DNA((I)和(II))的非共价相互作用显著地激活，因此在生理条件下金属离子的位置选择性断裂迅速进行。序列选择性的RNA断裂也是通过稀土(III)离子和一个在其内部位置含有吖啶的DNA的非共价组合(二元系)实现的。目标磷酸二酯键也被修饰的DNA激活，并优先被水解。DNA-ACR/Lu(III)二元体系仍然比三元体系具有更高的活性(约20%)。

Non-enzymatic hydrolysis of DNA has been one of the most significant targets for chemists. Several years ago, the remarkable catalysis by the lanthanide ions was found, and highly stable phosphodiester linkages in DNA were hydrolyzed at pH 7. The Ce (IV) ion is especially active. However, the Ce (IV) easily forms a gel of metal hydroxide, and this feature imposes limitations to the scope of its application. Homogeneous and catalytically active Ce (IV) complexes are required for more versatile applications. In this research, we fourcused on the development of (1) catalytically active Ce (IV) complexes, and (2) the methods of the activation of the target phosphodiesters in nucleic acids.(1) Homogeneous Ce (IV) complex of EDTA promptly hydrolyzes oligonucleotides under physiological conditions. In contrast, dinucleotides are not hydrolyzed to measurable extents. The degree of polymerization of DNA substrate is crucial for the present catalysis. The activity of this complex is significantly increased by the cooperation with selected oligoamines. The reaction in the presence of spermine (10 μmol dm^<-3>) is about 5O times as fast as that in its absence.(2) Non-covalent systems (ternary system), composed of (i) DNA oligomer bearing an acridine, (ii) unmodified DNA, and (iii) free lanthanide (III) ion, selectively and efficiently hydrolyse RNA at the target pohosphodiester. The corresponding phosphodiester linkage is notably activated by non-covalent interactions with the DNAs ((i) and (ii)), and thus the site-selective scission by the metal ion promptly proceeds under physiological conditions. The sequence-selective RNA scission is also achieved by non-covalent combinations of lanthanide (III) ion and one DNA bearing an acridine in its intern al position (binary system). The target phosphodiester linkage is also activated by the modified DNA, and preferentially hydrolyzed. The DNA-Acr/Lu (III) binary system is still more (about 20%) active than the ternary system.