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Petra Wallenmeyer MS Thesis.pdf (1.63 MB)

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Investigating Current Mechanistic Models of DNA Replication and Repair

Wallenmeyer, Petra C, Wallenmeyer
http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662

Abstract Details

2017, Master of Science, Ohio State University, Chemistry.
DNA polymerases (pols) play a pivotal role in both the replication and the repair of genomic DNA. Replicative pols are highly accurate and processive, synthesizing long stretches of DNA in a single binding event, while repair and bypass pols are error-prone and only able to incorporate a few nucleotides before dissociation. During replication, the pol may encounter DNA modifications induced by endogenous and exogenous factors such as oxidative metabolites, UV radiation, or epigenetic additions. These modifications may alter the local structure of DNA, resulting in inhibition of the replicative pol and stalling of the replication machinery. When the replicative pol stalls, a repair or bypass pol can take over and perform translesion synthesis (TLS). During TLS, a nucleotide is inserted opposite a lesion on the template DNA strand before the replicative pol can continue DNA synthesis. The mechanistic details of DNA replication, bypass, and repair are areas of ongoing research and are important to other areas of research such as drug design, cancer research, metabolism, and aging. The overarching goal of my research was to contribute to the mechanistic understanding of how pols perform DNA synthesis and bypass of DNA lesions. With this goal in mind, one of my main projects was to investigate the bypass kinetics of a common epigenetic signal, modification of the C5-position on cytosine (5xC). I used a specialized pol, human pol iota, to conduct this investigation. Using pre-steady state kinetic methods, I determined the dissociation constant (Kd) and maximum incorporation rate (kpol) for each deoxynucleoside (dNTP) opposite each 5xC modification. I also attempted to determine the structural details of the two accessory subunits of human pol epsilon, a replicative DNA pol that carries out leading strand synthesis, via X-ray crystallography to propose a hypothesis for its intrinsically high fidelity and processivity. Other projects I was involved in included investigating the biochemical properties of human PrimPol, the second pol found to operate in the mitochondria, using pre-steady state kinetics; using X-ray crystallography and pre-steady state kinetics to characterize human pol beta, and determining the best assay conditions to perform pre-steady state kinetics experiments with Zika Virus NS5, an RNA-dependent RNA polymerase that replicates viral RNA genome.
Zucai Suo, Dr. (Advisor)
James Cowan, Dr. (Committee Member)
Thomas Magliery, Dr. (Committee Member)
107 p.
Biochemistry; Chemistry
DNA polymerases; DNA repair; DNA replication; epigenetics; modified cytosine; x-ray crystallography; pre-steady state kinetics

Recommended Citations

Citations

  • Wallenmeyer, Wallenmeyer, P. C. (2017). Investigating Current Mechanistic Models of DNA Replication and Repair [Master's thesis, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662

    APA Style (7th edition)

  • Wallenmeyer, Wallenmeyer, Petra. Investigating Current Mechanistic Models of DNA Replication and Repair. 2017. Ohio State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662.

    MLA Style (8th edition)

  • Wallenmeyer, Wallenmeyer, Petra. "Investigating Current Mechanistic Models of DNA Replication and Repair." Master's thesis, Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662

    Chicago Manual of Style (17th edition)

osu1503321742475662
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