Cisplatin (cis-Pt(NH3)2Cl2) and cis-[Pt(NH3)2(H2O)2]2+ Intrastrand Cross-Linking Reactions at the Telomere GGGT DNA Sequence Embedded in a Duplex, a Hairpin, and a Bulged Duplex:  Use of Mg2+ and Zn2+ to Convert a Hairpin to a Bulged Duplex

• Published in: Inorganic chemistry Vol. 38; no. 26; pp. 6069 - 6080
• Main Authors: Villanueva, Julie M, Jia, Xin, Yohannes, Paulos G, Doetsch, Paul W, Marzilli, Luigi G
• Format: Journal Article
• Language: English
• Published: American Chemical Society 27-12-1999
• ISSN: 0020-1669; 1520-510X
• DOI: 10.1021/ic990603f

In the past, we showed that metal species have a high affinity for the central G in the GGG sequence of the duplex d(A1T2G3G4G5T6A7C8C9C10A11T12)2 (G3-D) and that cisplatin (cis-Pt(NH3)2Cl2) and G3-D formed an N7−Pt−N7 G4,G5 intrastrand cross-link preferentially over the G3,G4 adduct (∼25:1). Thus, a putative G4 monoadduct was postulated to cross-link in the 3‘- rather than the normally more favorable 5‘-direction. To evaluate this hypothesis and also to explore why the G3-D G4,G5 adduct had an unusual hairpin structure, we have now introduced the use of N,N‘-dimethylthiourea (DMTU) as a monoadduct trap and have extended the study to a G3-D analogue with a hairpin form, d(A1T2G3G4G5T6T7C8C9C10A11T12) (G3-H). Chemical shift and 2D 1H and 13C NMR data indicated that the G3-H hairpin has a stem region with B-form structure and a nonhelical loop region. Zn2+ or Mg2+ ions transformed G3-H into a bulged duplex. Downfield shifts of G4H8 and G4C8 NMR signals indicated that Zn2+ binds preferentially to G4N7. Reaction of cisplatin or cis-[Pt(NH3)2(H2O)2]2+ with the bulged duplex and hairpin forms of G3-H gave a similar intrastrand cross-link ratio, G4,G5:G3,G4 = 7:3. This ratio is insensitive to DNA form or Pt leaving group. For G3-D this ratio is lower in the cis-[Pt(NH3)2(H2O)2]2+ reaction (∼1:1) than in the cisplatin reaction (25:1), indicating that the leaving group influences the cross-linking step for G3-D. The G4 monoadducts of the cis-Pt(NH3)2Cl2−G3-H and −G3-D and the cis-[Pt(NH3)2(H2O)2]2+−G3-D reactions were trapped with DMTU, but no monoadduct was trapped in the cis-[Pt(NH3)2(H2O)2]2+−G3-H reaction. The results suggest that the respective monoadducts are more long-lived for G3-D. We postulate that the G5 in the G3-D Cl-G4 monoadduct is placed in a favorable position to form the cross-link because of a prior conformational change induced by G4−A7 stacking. This accounts for the very high selectivity for 3‘-cross-linking. Nevertheless, in all other cases, regardless of the form or conformation, 3‘-direction cross-linking is unusually favored at GGGT sequences, suggesting that the sequence itself contributes greatly to the 3‘-cross-linking preference; since telomeres have multiple repeats of this GGGT sequence, this finding may have biological relevance.