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Title: Decoding the structure WWP2 in the TGFβ signalling pathway
Author: Wahl, Lloyd
ISNI:       0000 0004 5916 0471
Awarding Body: University of East Anglia
Current Institution: University of East Anglia
Date of Award: 2016
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The secret to the specificity of the ubiquitin-proteasome system lies in the protein-protein interaction domains of the diverse group of E3 enzymes. WWP2 is one such E3 enzyme, and the relevant protein-protein interaction domain is the WW domain. WWP2 has four WW domains which are used to interact with proline-rich motifs found in the sequences of the Smad signalling proteins that propagate or inhibit the TGFβ pathway, and in so doing, allows WWP2 to regulate its Smad targets. WWP2 has three isoforms that are known to participate in regulation of TGFβ signalling, but, even amongst isoforms of the same E3, they exhibit different specificities for components of the pathway. The reason for this is unknown, but it is likely to be due to the different domain composition of WWP2, since each of the isoforms has a different combination of WW domains. The aim of this thesis is to investigate the structure of the domains of WWP2, and to explore how this relates to the selectivity of different isoforms in the TGFβ pathway. Overexpression of recombinant WWP2 domains in a bacterial host, and affinity and sizeexclusion chromatography have been used to produce pure, high concentration protein samples. Both NMR spectroscopy and crystallography have been used in an attempt to elucidate the structure of WWP2 domains. NMR spectroscopy, the more successful of the two approaches, has allowed the elucidation of the structure of the fourth WW domain of WWP2. By observing ligand interaction using NMR, the binding site of WW4 is revealed and the substrate preference of WW4 and WW3 domains is observed on a molecular level. Evidence of phospho-regulation of substrate selectivity is presented, and a structural basis for this selectivity is proposed. In addition, a further layer of complexity is added to the WWP2 isoform-mediated regulation of the TGFβ signalling pathway, as a new isoform is discovered.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available