Competitive formation of DNA linkage isomers by a trinuclear platinum complex and the influence of pre-association

Abstract

2D [¹H, ¹⁵N] HSQC NMR spectroscopy has been used to monitor the reaction of fully ¹⁵N-labelled [{trans-PtCl(NH₃)₂}₂(μ-trans-Pt(NH₃)₂{NH₂(CH₂)₆NH₂}₂)]⁴⁺ (BBR3464 (¹⁵N-1)) with the 14-mer duplex (5′-{d(ATACATG(7)G(8)TACATA)}-3′·5′-{d(TATG(18)TACCATG(25)TAT)}-3′ or I) at pH 5.4 and 298 K, to examine the possible formation of 1,4 and 1,5-GG adducts in both 5′-5′ and 3′-3′ directions. In a previous study, the binding of the dinuclear 1,1/t,t to I showed specific formation of the 5′-5′ 1,4 G(8)G(18) cross-link, whereas in this case a mixture of adducts were formed. Initial ¹H NMR spectra suggested the presence of two pre-associated states aligned in both directions along the DNA. The pre-association was studied in the absence of covalent binding, by use of the “non-covalent” analog [{trans-Pt(NH₃)₃}₂(μ-trans-Pt(NH₃)₂{NH₂(CH₂)₆NH₂}₂)]⁶⁺ (AH44, 0). Chemical shift changes of DNA protons combined with NOE connectivities between CH₂ and NH₃ protons of 0 and the adenine H2 protons on I show that two different molecules of 0 are bound in the minor groove. Molecular dynamic simulations were performed to study the interaction of 0 at the two pre-association sites using charges derived from density functional theory (DFT) calculations. Structures where the central platinum is located in the minor groove and the aliphatic linkers extend into the major groove, in opposite directions, often represent the lowest energy structures of the snapshots selected. In the reaction of ¹⁵N-1 and I, following the pre-association step, aquation occurs to give the mono aqua monochloro species 2, with a rate constant of 3.43 ± 0.03 × 10⁻⁵ s⁻¹. There was evidence for two monofunctional adducts (3, 4) bound to the 3′ (G8) and 5′ (G7) residues and the asymmetry of the ¹H,¹⁵N peak for 3 suggested two conformers of the 3′ adduct, aligned in different directions along the DNA. The rate constant for combined monofunctional adduct formation (0.6 ± 0.1 M⁻¹) is ca. 2-fold lower for 1 compared to 1,1/t,t, whereas the rate constant for conversion of the combined monofunctional species to combined bifunctional adducts (5) (8.0 ± 0.2 × 10⁻⁵ s⁻¹) is two-fold higher.