Investigating Current Mechanistic Models of DNA Replication and Repair

Investigating Current Mechanistic Models of DNA Replication and Repair

Bibliographic Details
Main Author: Wallenmeyer, Petra C., Wallenmeyer
Language:English
Published: The Ohio State University / OhioLINK 2017
Subjects:
Biochemistry
Chemistry
DNA polymerases
DNA repair
DNA replication
epigenetics
modified cytosine
x-ray crystallography
pre-steady state kinetics
Online Access:http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662
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spelling ndltd-OhioLink-oai-etd.ohiolink.edu-osu15033217424756622021-08-03T07:03:57Z Investigating Current Mechanistic Models of DNA Replication and Repair Wallenmeyer, Petra C., Wallenmeyer Biochemistry Chemistry DNA polymerases DNA repair DNA replication epigenetics modified cytosine x-ray crystallography pre-steady state kinetics 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. 2017 English text The Ohio State University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662 http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662 unrestricted This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws.
collection NDLTD
language English
sources NDLTD
topic Biochemistry
Chemistry
DNA polymerases
DNA repair
DNA replication
epigenetics
modified cytosine
x-ray crystallography
pre-steady state kinetics
spellingShingle Biochemistry
Chemistry
DNA polymerases
DNA repair
DNA replication
epigenetics
modified cytosine
x-ray crystallography
pre-steady state kinetics
Wallenmeyer, Petra C., Wallenmeyer
Investigating Current Mechanistic Models of DNA Replication and Repair
author Wallenmeyer, Petra C., Wallenmeyer
author_facet Wallenmeyer, Petra C., Wallenmeyer
author_sort Wallenmeyer, Petra C., Wallenmeyer
title Investigating Current Mechanistic Models of DNA Replication and Repair
title_short Investigating Current Mechanistic Models of DNA Replication and Repair
title_full Investigating Current Mechanistic Models of DNA Replication and Repair
title_fullStr Investigating Current Mechanistic Models of DNA Replication and Repair
title_full_unstemmed Investigating Current Mechanistic Models of DNA Replication and Repair
title_sort investigating current mechanistic models of dna replication and repair
publisher The Ohio State University / OhioLINK
publishDate 2017
url http://rave.ohiolink.edu/etdc/view?acc_num=osu1503321742475662
work_keys_str_mv AT wallenmeyerpetracwallenmeyer investigatingcurrentmechanisticmodelsofdnareplicationandrepair
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