article published yes Romeo C. A. Dubini author Eva Korytiaková author Thea Schinkel author Pia Heinrichs author Thomas Carell author Petra Rovo author c316e53f-b965-11eb-b128-bb26acc59c000000-0001-8729-7326 NMR department IST Austria Open Access Fund project 5-Carboxycytosine (5caC) is a rare epigenetic modification found in nucleic acids of all domains of life. Despite its sparse genomic abundance, 5caC is presumed to play essential regulatory roles in transcription, maintenance and base-excision processes in DNA. In this work, we utilize nuclear magnetic resonance (NMR) spectroscopy to address the effects of 5caC incorporation into canonical DNA strands at multiple pH and temperature conditions. Our results demonstrate that 5caC has a pH-dependent global destabilizing and a base-pair mobility enhancing local impact on dsDNA, albeit without any detectable influence on the ground-state B-DNA structure. Measurement of hybridization thermodynamics and kinetics of 5caC-bearing DNA duplexes highlighted how acidic environment (pH 5.8 and 4.7) destabilizes the double-stranded structure by ∼10–20 kJ mol–1 at 37 °C when compared to the same sample at neutral pH. Protonation of 5caC results in a lower activation energy for the dissociation process and a higher barrier for annealing. Studies on conformational exchange on the microsecond time scale regime revealed a sharply localized base-pair motion involving exclusively the modified site and its immediate surroundings. By direct comparison with canonical and 5-formylcytosine (5fC)-edited strands, we were able to address the impact of the two most oxidized naturally occurring cytosine derivatives in the genome. These insights on 5caC’s subtle sensitivity to acidic pH contribute to the long-standing questions of its capacity as a substrate in base excision repair processes and its purpose as an independent, stable epigenetic mark. https://research-explorer.ista.ac.at/download/10758/11692/2022_ACSPhysChemAU_Dubini.pdf application/pdfno American Chemical Society2022 eng 2694-2445 3563778110.1021/acsphyschemau.1c00050 23237-246 https://www.biorxiv.org/content/10.1101/2021.12.14.472563 Dubini, R. C. A., Korytiaková, E., Schinkel, T., Heinrichs, P., Carell, T., &#38; Rovo, P. (2022). 1H NMR chemical exchange techniques reveal local and global effects of oxidized cytosine derivatives. <i>ACS Physical Chemistry Au</i>. American Chemical Society. <a href="https://doi.org/10.1021/acsphyschemau.1c00050">https://doi.org/10.1021/acsphyschemau.1c00050</a> Dubini RCA, Korytiaková E, Schinkel T, Heinrichs P, Carell T, Rovo P. 1H NMR chemical exchange techniques reveal local and global effects of oxidized cytosine derivatives. <i>ACS Physical Chemistry Au</i>. 2022;2(3):237-246. doi:<a href="https://doi.org/10.1021/acsphyschemau.1c00050">10.1021/acsphyschemau.1c00050</a> R.C.A. Dubini, E. Korytiaková, T. Schinkel, P. Heinrichs, T. Carell, P. Rovo, ACS Physical Chemistry Au 2 (2022) 237–246. Dubini, Romeo C. A., et al. “1H NMR Chemical Exchange Techniques Reveal Local and Global Effects of Oxidized Cytosine Derivatives.” <i>ACS Physical Chemistry Au</i>, vol. 2, no. 3, American Chemical Society, 2022, pp. 237–46, doi:<a href="https://doi.org/10.1021/acsphyschemau.1c00050">10.1021/acsphyschemau.1c00050</a>. Dubini RCA, Korytiaková E, Schinkel T, Heinrichs P, Carell T, Rovo P. 2022. 1H NMR chemical exchange techniques reveal local and global effects of oxidized cytosine derivatives. ACS Physical Chemistry Au. 2(3), 237–246. Dubini, Romeo C. A., Eva Korytiaková, Thea Schinkel, Pia Heinrichs, Thomas Carell, and Petra Rovo. “1H NMR Chemical Exchange Techniques Reveal Local and Global Effects of Oxidized Cytosine Derivatives.” <i>ACS Physical Chemistry Au</i>. American Chemical Society, 2022. <a href="https://doi.org/10.1021/acsphyschemau.1c00050">https://doi.org/10.1021/acsphyschemau.1c00050</a>. R. C. A. Dubini, E. Korytiaková, T. Schinkel, P. Heinrichs, T. Carell, and P. Rovo, “1H NMR chemical exchange techniques reveal local and global effects of oxidized cytosine derivatives,” <i>ACS Physical Chemistry Au</i>, vol. 2, no. 3. American Chemical Society, pp. 237–246, 2022. 107582022-02-16T11:18:21Z2025-04-15T06:53:09Z