Walker, Jennifer L
(2024)
A novel mutation in yeast Hxk2 confers 2DG resistance through altered dynamics of the active site.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
High rates of glycolysis are present both in cancer cells and yeast cultures. Using lab evolution with a glucose analog (2-deoxy-glucose or 2DG) and the drug-sensitive ABC16-monster yeast strain, I determined a probable cause of 2DG resistance to be a novel mutation in Hexokinase II (Hxk2G238V) that may alter the flexibility of amino acids outside of the hexose binding pocket. To verify that this mutation could possibly be the primary contributor to 2DG resistance found in my lab evolution experiments, we ruled out protein misfolding, protein under-expression, genomic abnormalities, and differences in gene copy number. We modeled the Hxk2G238V protein using molecular dynamics (MD) simulations, and I analyzed the simulations to find a region of increased flexibility lining the enzyme active site that is stabilized upon ligand binding. I describe the different opened and closed states that occurred in the wildtype and mutant-protein simulations and show that the Hxk2G238V protein has altered dynamics/movements that could explain some of the differences between the mutant and wildtype forms. Differences in flexibility in the ATP binding site in Hxk2G238V (versus wildtype) could be a main contributor to 2DG resistance, and further study may provide new insights about hexokinase function.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
19 December 2024 |
| Date Type: |
Publication |
| Defense Date: |
22 March 2022 |
| Approval Date: |
19 December 2024 |
| Submission Date: |
5 December 2024 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
139 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Biological Sciences |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Hxk2, yeast, hexokinase, ABC16 monster, molecular dynamics, lab evolution |
| Date Deposited: |
19 Dec 2024 21:12 |
| Last Modified: |
19 Dec 2024 21:12 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/42280 |
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