γ-Hydroxy-1, N 2-propano-2′-deoxyguanosine DNA adduct conjugates the N-terminal amine of the KWKK peptide via a carbinolamine linkage

Hai Huang, Hao Wang, Markus W. Voehler, Albena Kozekova, Carmelo J. Rizzo, Amanda McCullough, Robert (Stephen) Lloyd, Michael P. Stone

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Abstract

The γ-hydroxy-1,N 2-propano-2′-deoxyguanosine adduct (γ-OH-PdG) was introduced into 5′-d(GCTAGCXAGTCC)-3′ ·5′-d(GGACTCGCTAGC)-3′ (X = γ-OH-PdG). In the presence of excess peptide KWKK, 13C isotope-edited NMR revealed the formation of two spectroscopically distinct DNA-KWKK conjugates. These involved the reaction of the KWKK N-terminal amino group with the N 2-dG propylaldehyde tautomer of the γ-OH-PdG lesion. The guanine N1 base imino resonance at the site of conjugation was observed in isotope-edited 15N NMR experiments, suggesting that the conjugated guanine was inserted into the duplex and that the guanine imino proton was protected from exchange with water. The conjugates could be reduced in the presence of NaCNBH 3, suggesting that they existed, in part, as imine (Schiff base) linkages. However, 13C isotope-edited NMR failed to detect the imine linkages, suggesting that these KWKK conjugates existed predominantly as diastereomeric carbinolamines, in equilibrium with trace amounts of the imines. The structures of the diastereomeric DNA-KWKK conjugates were predicted from potential energy minimization of model structures derived from the refined structure of the fully reduced cross-link [Huang, H., Kozekov, I. D., Kozekova, A., Rizzo, C. J., McCullough, A., Lloyd, R. S., and Stone, M. P. (2010) Biochemistry, 49, 6155 -6164]. Molecular dynamics calculations carried out in explicit solvent suggested that the conjugate bearing the S-carbinolamine linkage was the major species due to its potential for intramolecular hydrogen bonding. These carbinolamine DNA-KWKK conjugates thermally stabilized duplex DNA. However, the DNA-KWKK conjugates were chemically reversible and dissociated when the DNA was denatured. In this 5′-CpX-3′ sequence, the DNA-KWKK conjugates slowly converted to interstrand N 2-dG:N 2-dG DNA cross-links and ring-opened γ-OH-PdG derivatives over a period of weeks.

Original languageEnglish (US)
Pages (from-to)1123-1133
Number of pages11
JournalChemical Research in Toxicology
Volume24
Issue number7
DOIs
StatePublished - Jul 18 2011

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Deoxyguanosine
DNA Adducts
Amines
Peptides
DNA
Imines
Guanine
Isotopes
Nuclear magnetic resonance
Bearings (structural)
Biochemistry
Schiff Bases
Molecular Dynamics Simulation
Hydrogen Bonding
Model structures
Potential energy
Molecular dynamics
Protons
Hydrogen bonds
Derivatives

ASJC Scopus subject areas

  • Toxicology

Cite this

γ-Hydroxy-1, N 2-propano-2′-deoxyguanosine DNA adduct conjugates the N-terminal amine of the KWKK peptide via a carbinolamine linkage. / Huang, Hai; Wang, Hao; Voehler, Markus W.; Kozekova, Albena; Rizzo, Carmelo J.; McCullough, Amanda; Lloyd, Robert (Stephen); Stone, Michael P.

In: Chemical Research in Toxicology, Vol. 24, No. 7, 18.07.2011, p. 1123-1133.

Research output: Contribution to journalArticle

Huang, Hai ; Wang, Hao ; Voehler, Markus W. ; Kozekova, Albena ; Rizzo, Carmelo J. ; McCullough, Amanda ; Lloyd, Robert (Stephen) ; Stone, Michael P. / γ-Hydroxy-1, N 2-propano-2′-deoxyguanosine DNA adduct conjugates the N-terminal amine of the KWKK peptide via a carbinolamine linkage. In: Chemical Research in Toxicology. 2011 ; Vol. 24, No. 7. pp. 1123-1133.
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abstract = "The γ-hydroxy-1,N 2-propano-2′-deoxyguanosine adduct (γ-OH-PdG) was introduced into 5′-d(GCTAGCXAGTCC)-3′ ·5′-d(GGACTCGCTAGC)-3′ (X = γ-OH-PdG). In the presence of excess peptide KWKK, 13C isotope-edited NMR revealed the formation of two spectroscopically distinct DNA-KWKK conjugates. These involved the reaction of the KWKK N-terminal amino group with the N 2-dG propylaldehyde tautomer of the γ-OH-PdG lesion. The guanine N1 base imino resonance at the site of conjugation was observed in isotope-edited 15N NMR experiments, suggesting that the conjugated guanine was inserted into the duplex and that the guanine imino proton was protected from exchange with water. The conjugates could be reduced in the presence of NaCNBH 3, suggesting that they existed, in part, as imine (Schiff base) linkages. However, 13C isotope-edited NMR failed to detect the imine linkages, suggesting that these KWKK conjugates existed predominantly as diastereomeric carbinolamines, in equilibrium with trace amounts of the imines. The structures of the diastereomeric DNA-KWKK conjugates were predicted from potential energy minimization of model structures derived from the refined structure of the fully reduced cross-link [Huang, H., Kozekov, I. D., Kozekova, A., Rizzo, C. J., McCullough, A., Lloyd, R. S., and Stone, M. P. (2010) Biochemistry, 49, 6155 -6164]. Molecular dynamics calculations carried out in explicit solvent suggested that the conjugate bearing the S-carbinolamine linkage was the major species due to its potential for intramolecular hydrogen bonding. These carbinolamine DNA-KWKK conjugates thermally stabilized duplex DNA. However, the DNA-KWKK conjugates were chemically reversible and dissociated when the DNA was denatured. In this 5′-CpX-3′ sequence, the DNA-KWKK conjugates slowly converted to interstrand N 2-dG:N 2-dG DNA cross-links and ring-opened γ-OH-PdG derivatives over a period of weeks.",
author = "Hai Huang and Hao Wang and Voehler, {Markus W.} and Albena Kozekova and Rizzo, {Carmelo J.} and Amanda McCullough and Lloyd, {Robert (Stephen)} and Stone, {Michael P.}",
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AU - Wang, Hao

AU - Voehler, Markus W.

AU - Kozekova, Albena

AU - Rizzo, Carmelo J.

AU - McCullough, Amanda

AU - Lloyd, Robert (Stephen)

AU - Stone, Michael P.

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