Femtosecond broadband time-resolved fluorescence and transient absorption spectroscopic study of excited states of guanine, G-quadruplex, and cytosine and its derivatives

Abstract

Ultraviolet (UV) irradiation of DNA produces electronic excited states that may lead to formation of harmful photo-products, e.g. cyclobutane pyrimidine dimer and 8-oxo-guanine derived respectively from cytosine (C/Cyt) and guanine (G) containing DNAs are linked to mutagenesis and skin cancer. Despite the large amount of studies, the precise nature and dynamics of excited states of G, C, and related DNA assemblies are far from fully understood. This thesis reports a direct experimental investigation of the excited states of three DNA systems which are (i) 2’-deoxyguanosine (dGuo), 2’-deoxyguanosine 5’-monophosphate (dGMP); (ii) self-assembled G-quadruplexes formed by human telomeric sequence (HTS) in the presence of sodium (HTS/Na+) or potassium cation (HTS/K+); and (iii) cytosine, its nucleosides, nucleotides and a range of selected C5-substituted derivatives including 5-methylcytosine, its nucleosides, and 5-fluorocytosine. The studies were conducted by employing a combination of femtosecond (fs) broadband time-resolved fluorescence (fs-TRF), fluorescence anisotropy (fs-TRFA), and fs transient absorption (fs-TA) coupled with steady-state spectroscopy performed with excitation at varied wavelengths and in solvents of different properties. The results obtained for dGuo and dGMP lead to identification of a new species that is attributed to a weakly emissive πσ* state with charge transfer character. The species is favored at higher excitation energy and formed within ~50 fs after the photo-excitation. The dynamics of πσ* state was found to vary with the solvent hydrogen bonding capacity. A distinct deactivation model mediated by the πσ* along with two ππ* states, the Laππ* and Lbππ*, is proposed to account for nonradiative decay of UV-excited dGuo and dGMP. The results obtained for HTS/Na+ and HTS/K+, and for the corresponding stoichiometric mixture of monomeric bases show co-existence in HTS G-quadruplexes of two species having ~8 and 100 ps lifetime that are not seen in the monomeric mixture. These weakly emissive species dominate the overall deactivation processes. The ~8 and 100 ps species are attributed respectively to ππ* excitonic state without and with significant charge transfer between the neighboring G-tetrad planes. K+ Compared to , the presence of Na+ favors formation of the latter state. The inter-base interactions in G-quadruplex including stacking and Hoogsteen hydrogen bonding play a governing role for the nature and dynamics of excited-state decay. The results obtained for C, its nucleosides, nucleotides, and C5-substituted derivatives in neutral water reveal direct and systematic evidence that the substitution can affect strongly the excited-state dynamics. While C, its nucleosides, and nucleotides feature decay processes mediated by the photo-prepared ππ* and a “dark natured nπ* state, the C5-substituted derivatives feature decay process mediated solely by the ππ* state. The dynamics of ππ* state is affected strongly by the substituent at C5-position. Changing solvent from water to methanol affects only subtly the lifetime of ππ* state but it increases the nπ* state lifetime by ~4-7 folds for C, its nucleosides, and nucleotides. The results obtained add important spectral and dynamic parameters for elucidating the photophysics and the photostability of nucleobases and DNAs. The data presented are essential for understanding the potential photodamage of DNAs, especially those containing G and C.