DNA sequence context and protein composition modulate HMG-domain protein recognition of ciplatin-modified DNA

Proteins containing the high mobility group (HMG) DNA-binding domain form specific complexes with cisplatin-modified DNA which shield the major intrastrand d(GpG) and d(ApG) crosslinks from excision repair. The molecular basis for the specificity of binding was investigated for the two isolated domains of HMG1 with a series of 15-bp oligonucleotides, d(CCTCTCN(1)G*G*N2TCTTC).(GAAGAN(3)CCN(4)GAGAGG), where asterisks denote N-7-modification of guanosine with cisplatin. Alteration of the nucleotides flanking the platinum lesion modulated HMG1domA recognition in this series by over 2 orders of magnitude and revealed an unprecedented preference for N-2 = dA > T > dC. The flanking nucleotide preference for HMG1domB interaction with this oligonucleotide series was less pronounced and had only a 20-fold range of binding affinities. For the N-1 = N-2 = dA 15-bp probe, 100-fold stronger binding occurred with HMG1domA (K-d = 1.6 +/- 0.2 nM) compared to HMG1domB (K-d = 134 +/- 18 nM). The platinum-dependent recognition of the N-1 = N-2 = dA 15-bp probe saturates at 1 equiv of HMG1domA and is highly specific, as evidenced by the 1000-fold decrease in HMG1domA binding affinity for the corresponding unplatinated oligonucleotide. HMG domains were unable to bind specifically to cisplatin-modified DNA-RNA hybrids, revealing the need for a deoxyribose sugar backbone for specific complex formation with HMG-domain proteins. Protein-DNA contacts which may account for these observed binding preferences are proposed, and potential implications for the biological processing of cisplatin-DNA adducts are discussed.