Tracking Isomerizations of High-Energy Adenine Cation Radicals by UV-Vis Action Spectroscopy and Cyclic Ion Mobility Mass Spectrometry
Abstract
We report experimental and computational studies of protonatedadenine C-8 σ-radicals that are presumed yet elusive reactiveintermediates of oxidative damage to nucleic acids. The radicals weregenerated in the gas phase by the collision-induced dissociation of C-8-Br and C-8-I bonds in protonated 8-bromo- and 8-iodoadenineas well as by 8-bromo- and 8-iodo-9-methyladenine.
Protonation byelectrospray of 8-bromo- and 8-iodoadenine was shown by cyclic-ionmobility mass spectrometry (c-IMS) to form the N-1-H, N-9-H and N-3-H,N-7-H protomers in 85:15 and 81:19 ratios, respectively, in accordancewith the equilibrium populations of these protomers in water-solvatedions that were calculated by density functional theory (DFT). Protonationof 8-halogenated 9-methyladenines yielded single N-1-H protomers,which was consistent with their thermodynamic stability.
The radicalsproduced from the 8-bromo and 8-iodo adenine cations were characterizedby UV-vis photodissociation action spectroscopy (UVPD) andc-IMS. UVPD revealed the formation of C-8 σ-radicals along withN-3-H, N-7-H-adenine pi-radicals that arose as secondary productsby hydrogen atom migrations.
The isomers were identified by matchingtheir action spectra against the calculated vibronic absorption spectra.Deuterium isotope effects were found to slow the isomerization andincrease the population of C-8 σ-radicals. The adenine cationradicals were separated by c-IMS and identified by their collisioncross sections, which were measured relative to the canonical N-9-Hadenine cation radical that was cogenerated in situ as an internalstandard.
Ab initio CCSD-(T)/CBS calculations of isomer energies showedthat the adenine C-8 σ-radicals were local energy minima withrelative energies at 76-79 kJ mol(-1) abovethat of the canonical adenine cation radical. Rice-Ramsperger-Kassel-Marcuscalculations of unimolecular rate constants for hydrogen and deuteriummigrations resulting in exergonic isomerizations showed kinetic shiftsof 10-17 kJ mol(-1), stabilizing the C-8 σ-radicals.C-8 σ-radicals derived from N-1-protonated 9-methyladenine werealso thermodynamically unstable and readily isomerized upon formation.