Single base DNA lesions, such as T:G mismatches, are commonly fixed by base excision repair mechanisms. These mismatches come about from the deamination of 5-methylcytosine. The T:G msimatch was studied in the DNA sequence context, [5'-(dCGCGAATTCGCG)-3']2, comparing to the mismatched DNA [5'-(dCGTGAATTCGCG)-3']2 and [5'-(dCGCGAATTTGCG)-3']2. NMR spectroscopy is useful when determining the structure, thermodynamics, and kinetics of biological molecules such as a small sequences of DNA. The kinetic and thermodynamic properties were investigated by 31P 2D solution NMR. 2D spectra of each DNA sequence were taken at various temperatures and the 1D slices were obtained for each individual phosphate peak. The linewidths of the 1D spectra are then fit for each phosphate backbone peak. The linewidths as a function of temperature are fit to a curve to extract the activation energy ΔG‡ and therefore the rate constant of exchange, kex. The ΔG gives the equilibrium between the BI and BII states of the DNA backbone. The ΔG and ΔG‡ shows the chemical exchange rate kex and different temperatures allow for the determination of the enthalpy of activation ΔH‡. The kex shows the exchange rate between the BI and BII main conformational states of the DNA backbone. The combined thermodynamic and kinetic parameters offer valuable insight into the mechanism of mismatch detection.

Kinetics and thermodynamics of BI-BII conformational states in T:G mismatches

ACS Spring Meeting 2021

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