Date of Graduation
Master of Science in Chemistry
DNA, DNA damage, NMR spectroscopy, structural biology, dihydrouracil
Other Biochemistry, Biophysics, and Structural Biology | Physical Chemistry
The effects of the dihydrouracil lesion in DNA were studied using two dimensional NMR spectroscopy. The sequence used was based off of the Drew-Dickerson Dodecamer, with the cytosine in the three position replaced by a dihydrouracil. All of the nonexchangeable proton chemical shifts, with the exception of the H2, H5’, and H5’’, of the lesioned DNA were identified using NOESY spectra and then compared to the chemical shift values of the Drew Dickerson Dodecamer. The largest differences in chemical shifts were observed in the nucleotides neighboring the lesion, both within the strand and on the opposite strand. The imino exchangeable protons were also studied, and it was found that the lesion significantly decreases the local stability of double stranded DNA. The phosphorous backbone was also studied via HSQC and 1D Phosphorous NMR. The lesion was found to have a significant effect on the conformation of the phosphorous backbone of the DNA molecule. In the lesioned sequence, the 5’ and 3’ phosphates of the lesion site had a difference in % BII from the control sequence by 36.7 % and -27.2 %, respectively, at 25 ◦C. This trend was observed in all temperatures studied. In addition, the lesioned sequence also saw a large difference in the % BII between the 5’ and 3’ phosphates of the lesioned site. In the lesioned sequence, this difference was 85.2 % at 25 ◦C, compared to only 21.3 % in the control sequence. These results imply that changes in the phosphodiester backbone could be a potential area of indirect readout used by repair enzymes to identify the dihydrouracil lesion.
© Benjamin M. Boyd
Boyd, Benjamin M., "Effects of the Dihydrouracil Lesion on DNA Using 1H/31P 1D and 2D Solution NMR" (2019). MSU Graduate Theses. 3456.