Cisplatin Adducts on a GGG Sequence within a DNA Duplex Studied by NMR Spectroscopy and Molecular Dynamics Simulations

Abstract

The antitumor drug cisplatin(cis-[PtCl2(NH3)2]) reacts with cellular DNA to form GG intrastrand adducts between adjacent guanines as predominant lesions. GGG sites have been shown to be hotspots of platination. To study the structural perturbation induced by binding of cisplatin to two adjacent guanines of a GGG trinucleotide,we examined here the decanucleotide duplex d[(G1C2C3G*4 G*5 G6T7-C8G9C10).d(G11C12G13A14C15C16C17G18-G19C20)] (dsCG*G*G) intrastrand cross-linked at the G* guanines by cis-{Pt(NH3)2}2+ using NMR spectroscopy and molecular dynamics (MD) simulations.The NMR spectra of dsCG*G*G were found to be similar to those of previously characterized DNA duplexes cross-linked by cisplatin at apyG*G*X site (py=pyrimidine; X=C,T, A). This similarity of NMR spectra indicates that the base at the 3'-side of the G*G*-Pt cross-link does not affect the structure to a large extent. An unprecedented reversible isomerization between the duplex dsCG*G*G (bearing a G*4 G*5 -Pt chelate) and duplex dsGG*G*T (bearing a G*5 G*6 -Pt chelate)was observed, which yielded a 40:60 equilibrium between the two intrastrand GG-Pt cross-links. No formation of interstrand cross-links was observed.NMR spectroscopic data of dsCG*G*G indicated that the deoxyribose of the 5'-G* adopts an N-type conformation, and the cytidines C3, C15,and C16 have average phase angles intermediate between S and N. The NMR spectroscopic chemical shifts of dsGG*G*T showed some fundamental differences to those of pyG*G*-platinum adducts but were in agreement with the NMR spectra reported previously for the DNA duplexes crosslinked at an AG*G*C sequence by cisplatin or oxaliplatin. The presence of apurine instead of a pyrimidine at the 5'-side of the G*G* cross-link seems therefore to affect the structure of the XG* step significantly.

Cisplatin GG-crosslinks within single-stranded DNA: Origin of the preference for left-handed helicity

“…This is in agreement with experimental observations: In single-stranded Pt-XG*G* adducts, the nucleotide X 5′ to G*G* retains the canonical S pucker [8,27], whereas in duplexes, this nucleotide tends to repucker to N [5,29]. It has been suggested that the repuckering to N of the X sugar could be the consequence of the particular geometry of the XG* step within the duplex [27].…”

“…This can be seen in Fig. 6 where the platinated GG fragment of the L1Sε t model for TG*G* is superimposed with that of the time-averaged, energy-minimized model for the G*G*-cisplatinated duplex d(GCCG*G*GTCGC)-d(GCGACCCGGC) [29]. In Fig.…”

“…The free energies of these two interaction modes are similar [33] and it is difficult to weight the participation of each mode exactly. The AMBER force-field has been shown to have the tendency to favor the direct hydrogen-bonding mode [29]. However, a shift of the conformational equilibrium towards indirectly hydrogen-bonded structures would merely favor L1Sε − over L1Sε t (see Tab.…”

“…6. Stereoview of the cisplatinated GG dinucleotide fragments of the model L1S ε t of TG*G* (red, this work), and of the time-averaged, energy-minimized model of the duplex d(GCCG*G*GTCGC)-d(GCGACCCGGC) (blue, [29]). The two structures are superimposed so that their 3′-G* bases coincide.…”

“…A salient feature of cisplatin-DNA crosslinks are hydrogen bonds donated by the NH 3 ligands of platinum [5,29]. The N-coordination to platinum polarizes the N\H bonds and makes them excellent hydrogen bond donors.…”