Shriver, Kirsten
(2017)
Determination of Novel Gene Targets of TETS via Non-Canonical Protein and DNA Modifications and Selective Inhibition.
Master's Thesis, University of Pittsburgh.
(Unpublished)
Abstract
The Ten-Eleven translocation (TET) family of enzymes mediates the oxidation of DNA methylation, but the specific gene targets and the role of these enzymes in human development and disease are not well understood. The overall goals of this work are to profile TET-specific gene targets and to develop potent and specific small molecule inhibitors for TET family members. To accomplish these goals, a photo-crosslinking system and non-canonically modified DNA substrates will be developed as chemical tools to elucidate TET member-specific gene targets. Additionally, the development of specific, small molecule inhibitors will be explored. Photo-crosslinkable TET enzymes were developed, via incorporation of a photo-activable unnatural amino acid into the catalytic domain; three mutant enzymes were found to crosslink to methyl cytosine containing DNA substrates. This crosslinking system will be generalized across family members and applied in cells to identify novel gene targets of TET enzymes. DNA substrates with non-canonical modifications were tested in the development of a DNMT-TET tandem system to functionalize oxidation products. Ethyl, allyl, and propyl substituted DNA substrates were successfully oxidized by both wild type and mutant TET proteins. Potential inhibitor compounds were screened against the TET2 wild type enzyme, and a few compounds indicated slight inhibitory activity. A fragment based approach was employed to design potential linked inhibitor compounds, which will be synthesized and tested against TET enzymes.
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Details
| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
15 June 2017 |
| Date Type: |
Publication |
| Defense Date: |
22 November 2016 |
| Approval Date: |
15 June 2017 |
| Submission Date: |
11 April 2017 |
| Access Restriction: |
5 year -- Restrict access to University of Pittsburgh for a period of 5 years. |
| Number of Pages: |
46 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Chemistry |
| Degree: |
MS - Master of Science |
| Thesis Type: |
Master's Thesis |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
TET, photo-crosslinking |
| Date Deposited: |
16 Jun 2017 00:20 |
| Last Modified: |
15 Jun 2022 05:15 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/31417 |
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