DNA unwinding produced by site-specific intrastrand crosslinks of the antitumor drug cis-diamminedichloroplatinum(II)

Abstract

The DNA unwinding produced by specific adducts of the antitumor drug cis-diamminedichloroplatinum(II) has been quantitatively determined. Synthetic DNA duplex oligonucleotides of varying lengths with two base pair cohesive ends were synthesized and characterized that contained site-specific intrastrand N7-purine/N7-purine cross-links. Included are cis-[Pt(NH3)2[d(GpG)]], cis-[Pt(NH3)2(d(ApG)]], and cis-[Pt(NH3)2[d(GpTpG)]] adducts, respectively referred to as cis-GG, cis-AG, and cis-GTG. Local DNA distortions at the site of platination were amplified by polymerization of these monomers and quantitatively evaluated by using polyacrylamide gel electrophoresis. The extent of DNA unwinding was determined by systematically varying the interplatinum distance, or phasing, in polymers containing the adducts. The multimer that migrates most slowly gives the optimal phasing for cooperative bending, from which the degree of unwinding can be obtained. We find that the cis-GG and cis-AG adducts both unwind DNA by 13 degrees, while the cis-GTG adduct unwinds DNA by 23 degrees. In addition, experiments are presented that support previous studies revealing that a hinge joint forms at the sites of platination in DNA molecules containing trans-GTG adducts. On the basis of an analysis of the present and other published studies of site-specifically modified DNA, we propose that local duplex unwinding is a major determinant in the recognition of DNA damage by the Escherichia coli (A)BC excinuclease. In addition, local duplex unwinding of 13 degrees and bending by 35 degrees are shown to correlate well with the recognition of platinated DNA by a previously identified damage recognition protein (DRP) in human cells.