Detection of Proteins that Recognize Platinum‐modified DNA Using Gel Mobility Shift Assay

Using a gel mobility shift assay, we have identified proteins, in the nuclear extracts of a human lung cancer cell line, that recognize cis‐diamminedichloroplatinum(II) (cis‐DDP, CDDP)‐modified DNA. A 158‐base‐pair double‐stranded DNA fragment, derived from pBR322 plasmid DNA, was modified by either CDDP, tetrachloro(dl‐trans)‐1,2‐diaminocyclohexaneplatinum(IV) (tetraplatin) or trans‐DDP (the stereoisomer of CDDP and clinically ineffective). These platinum drug‐modified probes were incubated with nuclear extracts and analyzed by gel mobility shift assay. Proteins in the extracts selectively recognized the clinically active platinum‐modified DNA fragment. No binding to the trans‐DDP‐modified DNA fragment was observed. These proteins may play a role in the cytotoxicity or in a DNA repair process.