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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 preliminary stage. Herein, we choose i-motif, a unique C-quadruplex structure composed of hemi-protonated 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 × 10 s), tautomerization (8.8 × 10 s), and hydration (5.3 × 10 s) 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 significant roles 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.
PMID: 30624927 [PubMed - as supplied by publisher]