Femtosecond and nanosecond broadband time-resolved spectroscopic study on excited state dynamics of C-rich oligomers, polymeric and natural nucleic acids

Abstract

Carcinogenic mutation of DNA induced by UV irradiation has been reported to arise from both the rapidly forming exciton ππ* state (<1 ps) and the longer lived charge transfer state of the photo-excited DNA assembly. For the latter case, interbase interaction in DNA plays a pivotal role in governing the deactivation mechanism toward either the photostability or photodamage pathway. Studying how interbase interaction affecting the excited states dynamics of photo-excited DNAs is important for understanding the mechanism behind these photoreactions. In this study, excited state dynamics of various nucleic acids containing different base sequences and adopting different ground state structures resulted in various interbase interactions were investigated. These nucleic acids include, i-motif (quadruplex structure resulted from the self-assembling of cytosine rich strands), polymeric model DNA (polydA-polydT) and RNA (polyrA-polyrU) duplex, and natural DNA (Calf Thymus DNA) and RNA (Yeast RNA). Measurement was achieved by the joint application of steady-state and time-resolved spectroscopy. Steady-state measurement was conducted by the steady-state absorption, steady-state emission and circular dichroism spectroscopy. Time-resolved measurement was recorded by the femtosecond broadband time-resolved fluorescence (fs-TRF), nanosecond broadband time-resolved fluorescence (ns-TRF) and femtosecond broadband transient absorption (fs-TA) spectroscopy. Result obtained from comparative study of i-motif and single-stranded structure of homocytosine dC20 and cytosine rich human telomeric sequence at acidic and alkaline pH, respectively, exhibits a strong low energy emission occurring at > 400 nm in the i-motif. This is associated with multiple slow decay kinetics occurring at sub picosecond to nanoseconds timescale. Among them, the excited state species featuring lifetime at ~ 70 ps was found to be decayed by proton transfer, which was arisen from the charge transfer occurring within the hemi-protonated base pair. Excited state dynamics and spectral properties of i-motif were found to be affected by factors, such as the base sequence, the pH condition and excitation energy. In addition, the presence of i-motif from human telomeric sequence at physiological pH condition was identified from the emission study recorded at 300 nm excitation. This suggests i-motif may able to take some roles in biological application. Result recorded for the emission study of polydA-polydT and polyrA-polyrU in relative to the constituent nucleobase reveals that a much complicated excited state decay dynamics is presented in the polymeric duplexes. This contains two different kinds of charge transfer states with decay mediated by proton transfer occurring at a few picoseconds and ten nanoseconds, respectively. In addition, the long living monomer-like emission occurring at ~ 330 nm with decay lifetime at ~ 2 ns was observed in both polymeric duplexes. This is attributed to the delayed fluorescence from monomeric ππ* state, which is repopulated by the dark charge transfer state. The population of this long living monomer-like emission exhibits a strong influence by the ground state structure, which was shown by observing a smaller population in polyrA-polyrU than in polydA-polydT and it is further reduced under unstable structure induced by neat water environment. In case of Calf Thymus DNA and Yeast RNA, a generally similar excited state dynamics was observed in the two natural nucleic acid systems. Both of them exhibit long living monomer-like emission arising from the repopulation of ππ* state mediated by dark natured charge transfer state. However, subtle differences in both spectral and decay dynamics were observed. For example, comparing to Calf Thymus DNA, Yeast RNA exhibits a more important emission contribution from the charge transfer relevant states occurring at low energy region. Charge transfer species with lifetime of ~ 50 ps with decay mediated by proton transfer was observed in Calf Thymus DNA. This however was not occurred in Yeast RNA. These differences are attributed to the different base sequence and conformation adopted by the two natural DNA and RNA systems. Despite of these differences, the predominant fraction (> 95%) of the overall excited state population decays back to the ground state at before hundred picoseconds in both systems. Result presented in this thesis provides additional information on how excited states dynamics of various nucleic acid systems are affected by different interbase interactions arising from both the ground state structure and base sequence. This is important for gaining more understanding on the mechanism behind the photostability and photodamage of DNA.