Influence of an Exocyclic Guanine Adduct on the Thermal Stability, Conformation, and Melting Thermodynamics of a DNA Duplex

G. Plum Eric, Kenneth J. Breslauer, Arthur P. Grollman, Frances F. Johnson

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As part of an overall program to characterize the impact of mutagenic lesions on the physiochemical properties of DNA, we report here the results of a comparative spectroscopic study on pairs of DNA duplexes both with and without an exocyclic guanine lesion. Specifically, we have studied a family of four 13-mer duplexes of the form d(CGCATGYGTACGC)·d(GCGTACZCATGCG) in which Y is either the normal deoxyguanosine residue (G) or the exocyclic guanine adduct 1, N2-propanodeoxyguanosine (X), while Z is either deoxycytosine (C) or deoxyadenosine (A). Thus, the four duplexes studied, which can be designated by the identity of their central Y·Z base pair, are a Watson-Crick duplex (GC), a duplex with a central mismatch (GA), and two duplexes with exocyclic guanine lesions (X), that differ only by the base opposite the lesion (XC and XA). The data derived from our spectroscopic measurements on these four duplexes have allowed us to evaluate the influence of the exocyclic guanine lesion, as well as the base opposite the lesion, on the conformation, thermal stability, and melting energetics of the host DNA duplex. To be specific, our circular dichroism (CD) spectra show that the exocyclic guanine lesion induces alterations in the duplex structure, while our temperature-dependent optical measurements reveal that these lesioninduced structural alterations reduce the thermal stability, the transition enthalpy, and the transition free energy of the duplex. We also find that the lesioninduced thermal destabilization of the duplex (ΔTm) primarily results from the presence of the modified guanine residue, while being relatively insensitive to whether the base opposite the lesion is a C or an A residue (e.g., XA and XC have the same Tm values). In other words, the presence of the lesion makes the duplex thermal stability insensitive to the base opposite it, a result that contrasts with the corresponding unmodified duplexes GA and GC which exhibit substantially different Tm values. By contrast, we find that the presence of the lesion does not alter the influence of the cross-strand partner on the duplex differential thermodynamic stability, ΔΔG. In other words, the GC and GA duplexes and the XC and XA duplexes exhibit identical differences in free energy (ΔΔG). Thus, the lesion alters the differential thermal stability (ΔTm) but not the differential thermodynamic stability (ΔΔG) of duplexes with C and A cross-strand partners. Our CD spectra reveal an intriguing pH-dependent structural transition with an apparent pKa near neutrality for the XA duplex which is absent in the other three duplexes. Because A residues are inserted preferentially opposite X lesions in vivo, this result suggests that the biological repair system may be confronted with a mixture of two structures under physiological conditions. We also find the XA and XC duplexes to be thermodynamically similar. Consequently, the observed biological preference for insertion of A over C residues across from the lesion cannot be rationalized simply in terms of thermodynamic differences between the final duplex states. Finally, our data reveal the thermodynamic consequences of the pH-induced, structural alterations in the XA duplex observed by NMR (Kouchakdjian etal., 1989, 1990) to be minimal. This surprising result suggests that DNA may be sufficiently plastic so as to accommodate significant structural alterations in a compensatory manner so as to minimize the net energetic cost.

Original languageEnglish (US)
Pages (from-to)12096-12102
Number of pages7
Issue number48
StatePublished - Feb 1 1992

All Science Journal Classification (ASJC) codes

  • Biochemistry


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