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dc.contributor.advisorBarran, Perditaen
dc.contributor.advisorHupp, Teden
dc.contributor.authorNicholson, Judithen
dc.date.accessioned2014-04-29T15:18:32Z
dc.date.available2014-04-29T15:18:32Z
dc.date.issued2011-11-23
dc.identifier.urihttp://hdl.handle.net/1842/8762
dc.description.abstractThe oncoprotein MDM2 has an integral role in cancer development via multiple signalling pathways. Two proteomic mass spectrometry screens, label-free with spectral counting quantitation and 8-plex iTRAQ were used to identify proteins up or downregulated over time by the MDM2 targeting drug Nutlin. A subset of previously identified MDM2 binding partners were identified as altered after Nutlin treatment, along with proteins which have not as yet been linked to MDM2 or p53. Proteins altered two hours after Nutlin treatment were screened for sequence similarity to an MDM2 binding consensus motif based on the BOX-I region of p53. Peptides corresponding to this motif were validated for MDM2 binding, and the mode of binding investigated using competition ELISA and thermal denaturation assays. Known MDM2 ligands such as Nutlin were shown to have a range of effects on the binding of these newly identified MDM2 peptides, which may be attributed to allosteric regulation of MDM2. The effects of Nutlin on two full length proteins identified by the MS screens, CypB and NPM, were confirmed in vivo. In vitro binding of MDM2 to CypB and PK, which contain BOX-I like motifs, was also demonstrated validating proteomic mass spectrometry screens as a method to identify new protein-protein interactions. To further investigate the potential of linear motifs to modulate protein-protein interactions, a peptide aptamer targeting the protein AGR2 was tested for effect on AGR2 and p53 in a cancer cell line.en
dc.language.isoen
dc.publisherThe University of Edinburghen
dc.relation.hasversionNicholson, J. & Hupp, T. R. The molecular dynamics of MDM2. Cell Cycle 9, 1878-81 (2010).en
dc.relation.hasversionFourtouna, A., Murray, E., Nicholson, J., Maslon, M. M., Pang, L. Y., Dryden, D. T. F., Hupp, T. R. The Anterior Gradient-2 Pathway as a Model for Developing Peptide-Aptamer Anti-Cancer Drug Leads that Stimulate p53 Function. Current Chemical Biology 3, 124-137 (2009).en
dc.relation.hasversionDastidar, S. G., Raghunathan, D., Nicholson, J., Hupp, T. R., Lane, D. P. & Verma, C. S. Chemical states of the N-terminal "lid" of MDM2 regulate p53 binding: Simulations reveal complexities of modulation. Cell Cycle 10, 82-9 (2011).en
dc.subjectMDM2en
dc.subjectProteomicsen
dc.subjectLinear motifsen
dc.subjectProtein-protein interactionsen
dc.titleProteomic investigation of the MDM2 interactome and linear motif interactionsen
dc.typeThesis or Dissertationen
dc.type.qualificationlevelDoctoralen
dc.type.qualificationnamePhD Doctor of Philosophyen


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