title: HMG-domain protein recognition of cisplatin 1,2-intrastrand d(GpG) cross-links in purine-rich sequence contexts.
abstract: HMG-domain proteins bind strongly to bent DNA structures, including cruciform and cisplatin-modified duplexes. Such protein-platinated DNA complexes, formed where the DNA is modified by the active cis but not the inactive trans isomer of diamminedichloroplatinum(II), are implicated in the cytotoxic mechanism of the drug. A series of oligonucleotide duplexes with deoxyguanosine nucleosides flanking a cis-[Pt(NH(3))(2)¿d(GpG)-N7(1),-N7(2)¿] cross-link have been synthesized. These probes were used to determine the flanking sequence dependence of the affinity of the individual HMG domains of HMG1 toward cisplatin-modified DNA. Nine related sequences, where N(1) and N(2) are not dG and GG is the 1,2-intrastrand cisplatin adduct in N(1)GGN(2), were previously investigated [Dunham, S. U., and Lippard, S. J. (1997) Biochemistry 36, 11428-11436]. Three of the seven remaining possible sequences for which N(1) and/or N(2) was dG were prepared here by using normal deoxyguanosine, but the rest, where N(1) is dG and N(2) is dA, dC, T, or dG, could not be isolated in pure form. These sequences were accessed by using the synthetic bases 7-deazaadenine and 7-deazaguanine, which lack the nucleophilic N7 atom in the purine ring. Deaza nucleotides accurately mimic the properties of the natural bases, allowing the interaction of the HMG-domain proteins with cisplatin-modified DNA to be examined. These experiments reveal that the flexibility of A.T versus G.C flanking base pairs, rather than base-specific contacts, determines HMG1domA protein selectivity. This conclusion was supported by use of mutant HMG1domA and HMG1domB proteins, which exhibit identical flanking sequence selectivity. The methods and results obtained here not only improve our understanding of how proteins might mediate cisplatin genotoxicity but also should apply more generally in the investigation of how other proteins interact with damaged DNA.
authors.affiliation: Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
authors.forename: S M
authors.initials: SM
authors.lastname: Cohen
authors.forename: Y
authors.initials: Y
authors.lastname: Mikata
authors.forename: Q
authors.initials: Q
authors.lastname: He
authors.forename: S J
authors.initials: SJ
authors.lastname: Lippard
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000378:metabolism; D001483:Base Sequence; D002945:Cisplatin / Q000737:chemistry* / Q000378:metabolism; D018736:DNA Adducts / Q000737:chemistry* / Q000378:metabolism; D015342:DNA Probes / Q000138:chemical synthesis / Q000378:metabolism; D003849:Deoxyguanosine / Q000737:chemistry* / Q000378:metabolism; D015226:Dinucleoside Phosphates / Q000737:chemistry* / Q000378:metabolism; D006147:Guanine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000378:metabolism; D006609:High Mobility Group Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009692:Nucleic Acid Heteroduplexes / Q000378:metabolism; D009841:Oligonucleotides / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D018736:DNA Adducts; D015342:DNA Probes; D015226:Dinucleoside Phosphates; D006609:High Mobility Group Proteins; D009692:Nucleic Acid Heteroduplexes; D009841:Oligonucleotides; D011994:Recombinant Proteins; C043837:7-deazaadenine; D006147:Guanine; D003849:Deoxyguanosine; C066856:7-deazaguanine; D000225:Adenine; D002945:Cisplatin