Photoreactivity of platinum(II) in cisplatin-modified DNA affords specific cross-links to HMG domain proteins

Cisplatin-modified DNA forms specific complexes with proteins that contain the DNA binding motif known as the high-mobility group (HMG) domain. As a tool for investigating the role of these proteins in mediating the cytotoxic effects of cisplatin, a set of cisplatin analogs was prepared in which one of the ammine ligands was replaced with a photoreactive tethered aryl azide ligand. The ability of DNA modified by these platinum complexes to photo-cross-link to HMG1 was investigated. During this study, it was discovered that DNA modified with cisplatin itself can undergo photoinduced cross-linking to HMG1 when irradiated with 300 nm light. The covalent complexes resulting from this latter crosslinking reaction are completely reversed by the addition of sodium cyanide and can be degraded by proteinase K. These results confirm the presence of a protein-DNA cross-link and demonstrate that the platinum atom itself forms the point of attachment. By contrast, DNA modified with trans-diamminedichloroplatinum(II), [Pt(dien)Cl]Cl, or [Pt(NH3)(3)Cl]Cl does not cross-link to HMG1 upon irradiation. The photochemistry was exploited to cross-link a 15-base pair oligonucleotide containing a single, site-specific cis-[Pt(NH3)(2){d(GpG)-N7(1),-N7(2)}] intrastrand adduct to domain B of HMG1. Following proteolytic digestion of the resulting covalent complex, the site of attachment to the protein was determined by Edman degradation of the resulting peptide-DNA complex to be a single residue on HMG domain B, Lys-6. The data further suggest that this amino acid binds to platinum at a site made available by photolabilization of a purine ligand. These results afford the first structural information about the interaction of HMG domain proteins with cisplatin-modified DNA.