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Title: Semi-rational design and characterisation of peptide ligands to manipulate ID protein function
Author: Salman, Asma
Awarding Body: University of Essex
Current Institution: University of Essex
Date of Award: 2013
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ID helix-loop-helix (HLH) proteins play a central role in the regulation of mammalian cell growth, differentiation and tumourigenesis. In many human cancer types, the expression of one or more of the four 10 family members (ID1 to ID4) is deregulated. Extensive data supports a direct role for deregulated ID protein expression in conferring malignant properties on tumour cells, through their inhibition of basic HLH (bHLH) transcription factors, which highlights the ID proteins as candidate therapeutic targets. This research aimed to develop peptides capable of inhibiting ID protein function, and build on the current understanding of key residues within the HLH binding domain that govern the specificity of ID protein binding. Molecular modelling analysis facilitated the randomisation of several key residues at the HLH dimer interface of ID-binding transcription factor E47 to create peptide libraries. Using a semi-rational design approach, combined with a protein-fragment complementation screening strategy (PCA), two specific high-affinity ID1 and ID3-binding peptides were obtained, capable of blocking ID protein inhibition of E47. The ID proteins and their inhibitors were further characterised through thermodynamic analysis of their structure and stability using circular dichroism and a range of other biophysical techniques, and were found to display mostly helical structures that increased in stability on addition of PCA-selected inhibitors. The homo-and hetero-dimerisation of members of the ID family was also investigated, and complex size analysis revealed that 101, 3 and 4 can exist as tetrameric structures. In addition to this; heterodimeric interactions were observed between ID1 and the remaining ID proteins; the highest affinity of which was that between ID1 and ID4, which has not been previously reported. In future studies, ID peptide ligands can be explored for the development of anti-cancer and regenerative medicine therapeutics and used as experimental tools for dissecting molecular mechanisms of the ID proteins.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available