Synthesis Of Diiron(Ⅲ) Complexes And Their Activity In Cleavage Of DNA And Phosphodiesters

Mimicking the activities of nucleases is currently an attractive research area in molecularbiology since artificial nucleases have potential applications as novel restriction enzymes andanticancer therapeutic agents. In the past a few years, various synthetic metallonucleases havebeen reported as DNA hydrolytic agents. Nevertheless, their activity is still much lower thanthat of the corresponding enzymes. So the development of synthetic agents to cleave DNAwith high efficiency is a fascinating challenge.A new di-Fe(â…¢) complex (Fe₂L_b), linking a acridine group as DNA-intercalator, hasbeen designed and synthesized as hydrolytic cleaving agent of DNA. Compared with Fe₂L_a(without acridine group), Fe₂L_b has led to a 14-fold increase of the cleavage efficiency ofplasmid DNA as a result of the binding interaction between DNA and acridine moiety. Theinteraction has been demonstrated by UV-visible absorption, CD spectra, viscidityexperiment and thermal denaturation studies. The hydrolytic mechanism is supported by theevidence from T4 DNA ligase assay, reactive oxygen species (ROS) quenching experimentand DNA cleavage experiment under anaerobic condition.In nature, the high reactivity of nucleases is attributed to the cooperation of the protheticmetal irons with several functional groups of the amino acid side chains present in the activesite. The positively charged residues of lysine, arginine and histidine are thought to stabilizephosphorane-like transition states by electronstatic interactions, hydrogen bonding and/orproton transfer. Several examples of synthetic systems have been reported based on thesefunctions. Dramatically enhanced activity was observed for the systems containing hydrogenbonding donors. Based on L_a, we recently prepared two novel dinucleating ligands L_c and L_dwith bisammonium and bisguanidinium groups, respectively. The corresponding diironcomplexes Fe₂L_c and Fe₂L_d were obtained. The DNA cleavage experiment indicates a muchmore pronounced DNA cleavage efficiency for the introducing of bisammonium andbisguanidinium groups, and the cooperativities between metal ions and amino/guanidinegroups are evidenced by the fact that there is no significant cleavage after incubating DNAwith non-metallated ligands.The reactivity of DNA cleaving systems can be enhanced when their structures comprisegroups with high DNA affinity. As inspired by the strategy, a series of dinuclear ironcomplexes conjugating double-acridine, double-pyrene and double-anthracene moiety were synthesized. DNA cleavage activities were also investigated through incubating DNA in thepresence metal complexes. At the same conditions, the complex (Fe₂L_e) linked with doubleacridine groups introduces a remarkable scission of supercoiled DNA with a rate constant of1.2×10^-1 s^-1 at pH 7.0 and 37℃, and leads to a 170-fold increase of the cleavage efficiencywhen compared with a intercalator lacking diiron(â…¢) complex (Fe₂L_a). Furthermore, DNAbinding property experiments were carried out through absorption titration and result showsthe Fe₂L_e has also the highest DNA affinity.In order to verify the phophodiester cleavage activity of diiron(â…¢) complexes, a series ofkinetic experiments using BNPP and HPNP as substrates were carried out. The results showthe introduction of ammonium and guadinium groups can enhance the activity of diiron(â…¢)complex toward phosphodiester cleavage.