Degradation of Cytosine Radical Cations in 2′-Deoxycytidine and in i‑Motif DNA: Hydrogen-Bonding Guided Pathways

Radical cations of nucleobases are key intermediates causing genome mutation, among which cytosine C•+ is of growing importance because the ensuing cytosine oxidation causes GC → AT transversions in DNA replication. Although the chemistry and biology of steady-state C oxidation products have been characterized, time-resolved study of initial degradation pathways of C•+ is still at the preliminary stage. Herein, we choose i-motif, a unique C-quadruplex structure composed of hemiprotonated base pairs C­(H)+:C, to examine C•+ degradation in a DNA surrounding without interference of G bases. Comprehensive time-resolved spectroscopy were performed to track C•+ dynamics in i-motif and in free base dC. The competing pathways of deprotonation (1.4 × 107 s–1), tautomerization (8.8 × 104 s–1), and hydration (5.3 × 103 s–1) are differentiated, and their rate constants are determined for the first time, underlining the strong reactivity of C•+. Distinct pathway is observed in i-motif compared with dC, showing the prominent features of C•+ hydration forming C­(5OH)• and C­(6OH)•. By further experiments of pH-dependence, comparison with single strand, and with Ag+ mediated i-motif, the mechanisms of C•+ degradation in i-motif are disclosed. The hydrogen-bonding within C­(H)+:C plays a significant role in guiding the reaction flux, by blocking the tautomerization of C­(−H)• and reversing the equilibrium from C­(−H)• to C•+. The C radicals in i-motif thus retain more cation character, and are mainly subject to hydration leading to lesion products that can induce disruption of i-motif structure and affect its critical roles in gene-regulation.