| Artificial nuclease can mimic the function of natural nuclease by breaking the backbone of DNA site-specifically. And these small molecules can also be used as molecular biology tools and gene therapy agents. The present dissertation reported the design and synthesis of a series of artificial nuclease, and their detailed DNA binding, DNA cleavage and DNA condensation behaviors were also studied by many different experimental methods.In this paper, polyamides which have sequence-specificity to bases were chose as modificators and bis(2-benzimidazolylmethyl)amine (IDB), polyamines (Cyclen) with hydrolysis mechanism and ascorbic acid (Vc) with oxidative mechanism were chose as model compounds. Two parts above were linked by different flexible linker to form a series of new artificial nucleases. We expected that cleavage ability and sequence-specificity of small molecules were enhanced through the modification of polyamides. The cleavage agents mentioned above were synthesized and characterized.In the process of interaction research between cleavage agents and nucleic acid, gel electrophoresis experiences were performed to analysis the DNA cleavage discrepancy of three kinds of cleavage agents in the absence or presence of polyamide. The results showed that DNA cleavage ability of molecules modified by polyamide was stronger than that of original cleavage agents apparently in the same condition, and they can cleave duplex to form linear DNA through double strand cleavage in the lower concentration. Both hydrolytic (IDB& Cyclen) and oxidative (Vc) small cleavage agents are mediated efficiently by modification of polyamide moiety. In addition, statistic data suggested that the process of DNA cleavage resulted from Vc is completely random, but that of Vc modified by polyamide is selectivity and non-random.In the research of binding ability of cleavage agents and DNA, circular dichroism, DNA melting point, fluorescence quenching, UV spectrum and Maldi-Tof mass spectrometry were performed to quantify the affinity of cleavage agents in the absence and presence of polyamide to DNA. The data of binding constant, including C50,ΔTm and Kapp, showed that the interaction and affinity between cleavage agents and DNA increased apparently due to the introduction of polyamide. The binding mode has changed from electrostatic interaction to minor groove binding. The outstanding results further supported A-T rich sequences preference of cleavage agents in the presence of polyamide containing pyrrole units.An accident discovery happened in the research. Polyamine complexes containing imide can condense DNA to small particles under some condition. So, IDB, TACN and Cyclen were chose as model compounds and different side chains were linked, and nine polyamine ligands with different structures side chains were synthesized and characterized to discuss the effect of carbonyl to DNA condensation.The gel electrophoresis showed that polyamine with ortho-position carbonyl can condense DNA efficiently. However, the weak and none effect were obtained by the same ligand with meta-position carbonyl and without carbonyl, respectively. Zinc complex can condense pUC18 plasmid DNA to small particles which size is like about 1000bp band, but copper complex can cleave DNA while condense DNA to small particles with different sizes, showing smear band in the gel. Through the further experiments of concentration, temperature and time-dependence assay, we found that one of the necessary conditions is high temperature (50℃), and the other is higher concentration of reaction. The same assay was performed by choosing different kinds of DNA, and results demonstrated that the DNA condensation under some condition is general application.Atom force microscopy assay indicated that DNA condensation was centered on small molecules with positive charge, and DNA was condensed into the same or similar size particles with electrostatic interaction or hydrogen band.
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