Time-resolved Spectroscopy And Excited State Dynamics Of Protonated Adenosine And Cytosine Tetraplex Structures

There are three research systems in this paper:(1)protonated adenosine,(2)cytosine-rich human telomere sequence HTS and homologous oligonucleotide sequence d C20,(3)cytosinemethylated vascular endothelial growth factor(VEGF)promoter sequence,human RET oncogene promoter sequence and human telomere sequence(HTS).Utilizing circular dichroism,steady-state spectrum,femtosecond time-resolved fluorescence and absorption spectrum,combined with density functional and time-dependent density functional theoretical calculation,the following research results are obtained:1)The steady-state absorption and fluorescence spectra of the protonated adenosine were obtained,and the proton transfer in the excited state of the protonated adenosine was inferred from the comparison of the spectra.The dual exponential kinetic behavior of excited state decay of neutral(p H7)and protonated(p H3)adenosine was detected with different time constants,indicating that they have different excited state relaxation channels.The decay indexes of the excited state of protonated adenosine are 0.5ps and 2933 ps.Combined with experimental and calculational results,our study showed that the kinetic component of 0.5ps was derived from conical intersection internal conversion of excites state of syn-adenosine protonated at N3,while the long-lived 2933 ps was derived from excited state of syn-adenosine protonated at N1.Thus,adenosine protonation can significantly change the excited relaxation pathway.2)Under the experimental conditions,the human telomere cytosine-rich sequence HTS and the homologous oligonucleotide sequence d C20 folded into cytosine tetraplex structure.The steady-state absorption spectra indicated the structural differences between the two subjects.The femtosecond transient spectra revealed that base excited state and exciton state are the main transient species of excited state of HTS sequence tetraplex,while the long-lived exciplex is the main transient species of i-motif excited state of d C20 sequence tetraplex.The differences in the dynamics of the two excited states are mainly due to the base stacking degree and the number of bases in loop region.We performed kinetic analyses on the decay of the excitation spectra,and observed the multi-exponential kinetic behavior,which reflected the complexity of excited state relaxation of cytosine tetraplex structure,and that the excited state deactivation is dominated by the delocalization induced by stacking and base-pairing interactions.3)It was found that methylation of cytosine at different sites in sequence does not affect the formation of cytosine tetraplex structure,but has an important impact on its stability and its overall folding degree.With steady state spectra and corresponding fitting analyses,we found that methylation in the core region of the hemi-protonated base pairs,especially in the middle of the core region,can stabilize the i-motif structure to a greater extent,which is mainly showed in the regular variation of steady state fluorescence spectra,revealing the important role of the hemi-protonated base in the stability of DNA tetraplex structure.