Two-photon excitation two-dimensional fluorescence spectroscopy (2PE-2DFS) of the fluorescent nucleobase 6-MI

Claire S. Albrecht; Lawrence F. Scatena; Peter H. von Hippel; Andrew H. Marcus

doi:10.1117/12.3001802

13 March 2024 Two-photon excitation two-dimensional fluorescence spectroscopy (2PE-2DFS) of the fluorescent nucleobase 6-MI

Claire S. Albrecht, Lawrence F. Scatena, Peter H. von Hippel, Andrew H. Marcus

Author Affiliations +

Proceedings Volume 12863, Quantum Effects and Measurement Techniques in Biology and Biophotonics; 1286305 (2024) https://doi.org/10.1117/12.3001802
Event: SPIE BiOS, 2024, San Francisco, California, United States

Abstract

Base stacking is fundamentally important to the stability of double-stranded DNA. However, few experiments can directly probe the local conformations and conformational fluctuations of the DNA bases. Here we report a new spectroscopic approach to study the local conformations of DNA bases using the UV-absorbing fluorescent guanine analogue, 6-methyl isoxanthopterin (6-MI), which can be used as a site-specific probe to label DNA. In these experiments, we apply a two-photon excitation (2PE) approach to two-dimensional fluorescence spectroscopy (2DFS), which is a fluorescence-detected nonlinear Fourier transform spectroscopy. In 2DFS, a repeating sequence of four collinear laser pulses (with center wavelength ~ 675 nm and relative phases swept at radio frequencies) is used to excite the lowest energy electronic-vibrational (vibronic) transitions of 6-MI (with center wavelength ~ 340 nm). The ensuing low flux fluorescence is phase-synchronously detected at the level of individual photons and as a function of inter-pulse delay. The 2PE transition pathways that give rise to electronically excited state populations include optical coherences between electronic ground and excited states and non-resonant (one-photon-excited) virtual states. Our results indicate that 2PE-2DFS experiments can provide information about the electronic-vibrational spectrum of the 6-MI monomer, in addition to the conformation-dependent exciton coupling between adjacent 6-MI monomers within a (6-MI)₂ dimer. In principle, this approach can be used to determine the local base-stacking conformations of (6-MI)₂ dimer-substituted DNA constructs.

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(2024) Published by SPIE. Downloading of the abstract is permitted for personal use only.

Citation Download Citation

Claire S. Albrecht, Lawrence F. Scatena, Peter H. von Hippel, and Andrew H. Marcus "Two-photon excitation two-dimensional fluorescence spectroscopy (2PE-2DFS) of the fluorescent nucleobase 6-MI", Proc. SPIE 12863, Quantum Effects and Measurement Techniques in Biology and Biophotonics, 1286305 (13 March 2024); https://doi.org/10.1117/12.3001802

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