4.6 Article

F-19 electron nuclear double resonance (ENDOR) spectroscopy for distance measurements using trityl spin labels in DNA duplexes

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PHYSICAL CHEMISTRY CHEMICAL PHYSICS
卷 25, 期 35, 页码 23454-23466

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ROYAL SOC CHEMISTRY
DOI: 10.1039/d3cp02969g

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The combination of fluorine labeling and pulsed electron-nuclear double resonance (ENDOR) is a powerful technique for obtaining structural information about proteins and nucleic acids. This study explored the capability of Mims F-19 ENDOR experiments to report intermolecular distances in trityl- and F-19-labeled DNA duplexes. The results showed that the hyperfine splittings observed in the ENDOR spectra were consistent with the predicted interspin distances between F-19 and the trityl radical.
The combination of fluorine labeling and pulsed electron-nuclear double resonance (ENDOR) is emerging as a powerful technique for obtaining structural information about proteins and nucleic acids. In this work, we explored the capability of Mims F-19 ENDOR experiments on reporting intermolecular distances in trityl- and F-19-labeled DNA duplexes at three electron paramagnetic resonance (EPR) frequencies (34, 94, and 263 GHz). For spin labeling, we used the hydrophobic Finland trityl radical and hydrophilic OX063 trityl radical. Fluorine labels were introduced into two positions of a DNA oligonucleotide. The results indicated that hyperfine splittings visible in the ENDOR spectra are consistent with the most populated interspin distances between F-19 and the trityl radical predicted from molecular dynamic (MD) simulations. Moreover, for some cases, ENDOR spectral simulations based on MD results were able to reproduce the conformational distribution reflected in the experimental ENDOR line broadening. Additionally, MD simulations provided more detailed information about the melting of terminal base pairs of the oligonucleotides and about the configuration of the trityls relative to a DNA end.

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