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Characterizing the protonation state of cytosine in transient G·C Hoogsteen base pairs in duplex DNA.

Publication ,  Journal Article
Nikolova, EN; Goh, GB; Brooks, CL; Al-Hashimi, HM
Published in: J Am Chem Soc
May 8, 2013

G·C Hoogsteen base pairs can form transiently in duplex DNA and play important roles in DNA recognition, replication, and repair. G·C Hoogsteen base pairs are thought to be stabilized by protonation of cytosine N3, which affords a second key hydrogen bond, but experimental evidence for this is sparse because the proton cannot be directly visualized by X-ray crystallography and nuclear magnetic resonance spectroscopy. Here, we combine NMR and constant pH molecular dynamics simulations to directly investigate the pKa of cytosine N3 in a chemically trapped N1-methyl-G·C Hoogsteen base pair within duplex DNA. Analysis of NMR chemical shift perturbations and NOESY data as a function of pH revealed that cytosine deprotonation is coupled to a syn-to-anti transition in N1-methyl-G, which results in a distorted Watson-Crick geometry at pH >9. A four-state analysis of the pH titration profiles yields a lower bound pKa estimate of 7.2 ± 0.1 for the G·C Hoogsteen base pair, which is in good agreement with the pKa value (7.1 ± 0.1) calculated independently using constant pH MD simulations. Based on these results and pH-dependent NMR relaxation dispersion measurements, we estimate that under physiological pH (pH 7-8), G·C Hoogsteen base pairs in naked DNA have a population of 0.02-0.002%, as compared to 0.4% for A·T Hoogsteen base pairs, and likely exist primarily as protonated species.

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Published In

J Am Chem Soc

DOI

EISSN

1520-5126

Publication Date

May 8, 2013

Volume

135

Issue

18

Start / End Page

6766 / 6769

Location

United States

Related Subject Headings

  • Protons
  • Nucleic Acid Conformation
  • Molecular Dynamics Simulation
  • Magnetic Resonance Spectroscopy
  • General Chemistry
  • DNA
  • Cytosine
  • Crystallography, X-Ray
  • Base Pairing
  • 40 Engineering
 

Citation

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Nikolova, E. N., Goh, G. B., Brooks, C. L., & Al-Hashimi, H. M. (2013). Characterizing the protonation state of cytosine in transient G·C Hoogsteen base pairs in duplex DNA. J Am Chem Soc, 135(18), 6766–6769. https://doi.org/10.1021/ja400994e
Nikolova, Evgenia N., Garrett B. Goh, Charles L. Brooks, and Hashim M. Al-Hashimi. “Characterizing the protonation state of cytosine in transient G·C Hoogsteen base pairs in duplex DNA.J Am Chem Soc 135, no. 18 (May 8, 2013): 6766–69. https://doi.org/10.1021/ja400994e.
Nikolova EN, Goh GB, Brooks CL, Al-Hashimi HM. Characterizing the protonation state of cytosine in transient G·C Hoogsteen base pairs in duplex DNA. J Am Chem Soc. 2013 May 8;135(18):6766–9.
Nikolova, Evgenia N., et al. “Characterizing the protonation state of cytosine in transient G·C Hoogsteen base pairs in duplex DNA.J Am Chem Soc, vol. 135, no. 18, May 2013, pp. 6766–69. Pubmed, doi:10.1021/ja400994e.
Nikolova EN, Goh GB, Brooks CL, Al-Hashimi HM. Characterizing the protonation state of cytosine in transient G·C Hoogsteen base pairs in duplex DNA. J Am Chem Soc. 2013 May 8;135(18):6766–6769.
Journal cover image

Published In

J Am Chem Soc

DOI

EISSN

1520-5126

Publication Date

May 8, 2013

Volume

135

Issue

18

Start / End Page

6766 / 6769

Location

United States

Related Subject Headings

  • Protons
  • Nucleic Acid Conformation
  • Molecular Dynamics Simulation
  • Magnetic Resonance Spectroscopy
  • General Chemistry
  • DNA
  • Cytosine
  • Crystallography, X-Ray
  • Base Pairing
  • 40 Engineering