DNA cleavage behavior of a new p-xylyl spaced bisCu(BPA)Cl2 complex: the steric effect of a bulky p-xylyl-derived spacer

Abstract

The different DNA cleavage behaviors of 1,3- and 1,4-xylylene spaced bisCu(BPA)Cl₂ complexes (BPA, bis(pyridyl-2-methyl)amine) display the positioning effect of xylylene spacers, implying the essential roles of spacers in the related cleavage (Y. Zhao et al., Chem.–Eur. J., 2006, 12, 6621–6629). Herein, a p-xylyl derivative (2,1,3-benzoxadiazole-4,7-dimethylene (BDDM)) was utilized as a bulky spacer to construct a new p-xylyl spaced bisCu(BPA)Cl₂ complex 1 to explore the spacer's steric effect on the related DNA cleavage. Agarose gel electrophoresis demonstrated that complex 1 is able to convert the supercoiled plasmid pUC19 DNA into the nicked DNA and linear DNA in the presence of excessive reductant. This DNA cleavage behavior is different from that of simple p-xylylene spaced Cu₂(pTPXA)Cl₄, which mediates only the nicked DNA and random DNA fragments. The DNA binding ability of complex 1 is higher than that of Cu₂(pTPXA)Cl₄, while its DNA nicking ability is lower than that of Cu(BPA)Cl₂, as well as the dinuclear Cu₂(pTPXA)Cl₄, which displays a high DNA nicking efficiency due to the internuclear synergy in activating O₂ to damage DNA. The linear DNA formation induced by complex 1 is realized via a non-random double-stranded scission process. The temporal profiles of the nicked DNA and linear DNA in the presence of 8 μM complex 1 imply that the two sequential neighboring breaks in the complementary strands lead to the formation of linear DNA. With the DNA cleavage behavior in the presence of different reactive oxygen species (ROS) quenchers, the DNA-bound hydroxyl radical was proposed as the main ROS involved in the cleavage. Structural optimization of this complex disclosed its larger intramolecular Cu–Cu distance than Cu₂(pTPXA)Cl₄, which may disfavor the internuclear synergic O₂ activation. It was also proposed that the bulky BDDM spacer favors the two spaced copper centers function solely to damage in situ the two adjacent deoxyriboses from complementary strands to form the linear DNA, and the specific DNA binding mode of complex 1 due to the BDDM spacer disfavors the DNA-bound ROS to abstract a proton from deoxyribose and results in a lower DNA cleavage efficiency than mononuclear Cu(BPA)Cl₂. The non-random linear DNA mediating ability and the decreased DNA cleavage efficiency caused by the bulky internuclear spacer shown in the current study provides a new alternative to regulate the DNA cleavage behavior of polycopper artificial nucleases via spacer modification.