Fluorescence and Ultrafast Fluorescence Unveil the Formation, Folding Molecularity, and Excitation Dynamics of Homo‐Oligomeric and Human Telomeric i‐Motifs at Acidic and Neutral pH

Abstract

i-Motifs are tetraplex DNAs known to be stable at acidic pH. The structure of i-motifs is important in DNA nanotechnology; i-motif-forming sequences with consecutive cytosine (C) molecules are abundant throughout the human genome. There is, however, little information on the structure of C-rich DNAs under physiologically relevant neutral conditions. The electron dynamics of i-motifs, crucial to both biology and materials applications, also remains largely unexplored. In this work, we report a combined femtosecond and nanosecond broadband time-resolved fluorescence (TRF) and steady-state fluorescence investigation on homo-oligomer dC , a human telomeric sequence (HTS) 5'-dC (TA C ) , and its analogue performed with different excitation at both acidic and neutral pH. Our study provides direct observation of intrinsic fluorescence and the first full probe of the real-time dynamics of the intrinsic fluorescence from i-motifs formed from varied sequences and pH conditions. The results obtained demonstrate concrete evidence for the existence at neutral pH of i-motifs from both dC and the HTS. It also identifies that, under neutral conditions, the i-motif from dC adopting the bimolecular folding structure is significantly more stable than the HTS i-motif featuring the unimolecular topology. Our femtosecond and nanosecond TRF study unveils excitation dynamics distinctive of the interdigitated architecture of i-motifs with the excited states involved exhibiting deactivation over a remarkably broad timescale through multiple channels involving proton-coupled electron transfer lasting tens of picoseconds, as signified by the solvent kinetic isotope effect, and structure-dependent charge recombination in the hundreds of picoseconds to tens of nanoseconds time regime.

Real-time Monitoring Excitation Dynamics of Human Telomeric Guanine Quadruplexes: Effect of Folding Topology, Metal Cation, and Confinement by Nanocavity Water Pool

“…Although numerous researches have provided invaluable insights into, e.g., the thermodynamics, binding properties, and folding conformations of various GQs, , little is yet known about the excitation dynamics of GQs in general and the intramolecularly folded HT-GQs in particular. , Like nucleobases and other DNA secondary structures, GQs can absorb UV radiation strongly . Understanding the dynamics of their excited states created by UV absorption and factors impacting it is indispensable for elucidating at molecular level the photostability vs susceptibility to photodamage for GQs occurring in not only the telomeric region , but also in the promoter areas of many oncogenes. , Exploration of the excitation dynamics will also allow evaluating nature of the interbase electronic couplings distinctive of the GQs architecture, and providing information for improving application performance in materials science.…”

Hysteresis in poly‐2′‐deoxycytidine i‐motif folding is impacted by the method of analysis as well as loop and stem lengths

“…To reiterate, the present findings are important when researchers try to compare pH T values for a given sequence studied by different laboratories. [ 29,30 ]…”

Harnessing intrinsic fluorescence for typing of secondary structures of DNA

“…Over the years, the effects of guanine base orientation, size and nature of loops, presence and nature of bulges have been 100 investigated, contributing to establish some correlations between the G4 conformation and the emissive properties (29,31,32). Very recently, the investigation on intrinsic fluorescence has been extended to iM structures (33): upon excitation at 267 or 300 nm, iMs display a long-lived, broad emission band centered around 410-420 nm (Φ = 3.4-14 × 10 −4 , depending on the specific iM sequence, pH, and λex). The emission is postulated to arise 105 from the stacking of C:CH + base pairs from the two interpenetrated duplexes (33).…”

“…Very recently, the investigation on intrinsic fluorescence has been extended to iM structures (33): upon excitation at 267 or 300 nm, iMs display a long-lived, broad emission band centered around 410-420 nm (Φ = 3.4-14 × 10 −4 , depending on the specific iM sequence, pH, and λex). The emission is postulated to arise 105 from the stacking of C:CH + base pairs from the two interpenetrated duplexes (33).…”