White Rose University Consortium logo
University of Leeds logo University of Sheffield logo York University logo

Predicting and Testing Helix-Mimetic Inhibitors of the p53-Mdm2 Interaction

Dockray, Joel P (2015) Predicting and Testing Helix-Mimetic Inhibitors of the p53-Mdm2 Interaction. PhD thesis, University of Leeds.

Text (Thesis)
Dockray_JP_Biological_Sciences_PhD_2015.pdf - Final eThesis - complete (pdf)

Download (16Mb) | Preview


Aberrant protein-protein interactions (PPIs) are found in many disease states. Consequently, there is a need for PPI inhibitors for use as research tools and pharmaceutical lead compounds. Computational methods could greatly assist with the search for new PPIs. Oligobenzamides are novel PPI inhibitors which can theoretically be produced to display any sequence of side chains. Understanding the nature of oligobenzamide binding is important for identification of the most efficient strategy of predicting oligobenzamide inhibitors. The prediction of oligobenzamide affinities using thermodynamic integration and implicit solvent methods is described. Affinities of oligobenzamides for Mdm2 predicted using implicit solvent methods bore a moderate correlation with measured affinities. Examination of MM-PBSA results using analysis of variance revealed that it is not necessary to run simulations with every member of a large combinatorial library in order to predict their relative affinities because within a particular binding site, the degree of interaction between the side chains is small. However, it could be useful to separate molecules based on their predicted binding pose because oligobenzamides can bind to Mdm2 in many different ways, depending on the choice of side chains. This insight will be valuable for future attempts to predict oligobenzamide affinities. The 1H-15N HSQC NMR spectrum peaks of 15N-labelled Mdm2 L33E were assigned to facilitate the future validation of binding poses. An oligoamide was shown using NMR to bind in the correct place. However, NMR testing revealed that oligobenzamides can aggregate in aqueous solution despite being soluble. A novel FRET-based method was also developed which can be used to test potential inhibitors with a low solubility and high absorbance during their development. It was adapted for a microwell plate to facilitate future high throughput screening and an assay involving Cherry-labelled Mdm2 was tested which could be developed into an in vivo assay in the future.

Item Type: Thesis (PhD)
Keywords: protein-protein interactions, docking, molecular dynamics, MM-PBSA, thermodynamic integration, p53, Mdm2, oligobenzamide
Academic Units: The University of Leeds > Faculty of Biological Sciences (Leeds)
The University of Leeds > Faculty of Biological Sciences (Leeds) > Institute for Molecular and Cellular Biology (Leeds)
Identification Number/EthosID: uk.bl.ethos.651249
Depositing User: Dr Joel Dockray
Date Deposited: 22 Jun 2015 10:31
Last Modified: 11 Jul 2020 09:53
URI: http://etheses.whiterose.ac.uk/id/eprint/9201

You do not need to contact us to get a copy of this thesis. Please use the 'Download' link(s) above to get a copy.
You can contact us about this thesis. If you need to make a general enquiry, please see the Contact us page.

Actions (repository staff only: login required)