Intrinsically disordered proteins (IDPs) are proteins that lack well-defined three-dimensional structures and instead exist as dynamic ensembles of conformations. Post-translational modifications (PTMs), such as phosphorylation, are commonly used to regulate the function of IDPs by altering their electrostatic properties, secondary structure, and facilitating or preventing interactions with their binding partners. In recent years, IDPs have received a lot of attention due to their involvement in multiple neurodegenerative diseases and cancer. However, a mechanistic understanding of how modulation of charge interactions by phosphorylation affects the conformation-function relationships of IDPs is still lacking. We aim to contribute to the tools available for studying IDPs by first introducing PhosIDP, a web-tool developed to visualise the effects of phosphorylation in disordered regions. The effect of single, double, and triple phosphorylation on the helicity of the N-terminal transactivation domain (TAD) of p53 is explored by using three different molecular dynamics techniques. Our simulations demonstrate that upon single, double, and triple phosphorylation there is partial destabilisation of the helix around the phosphorylation sites, while there are only minor changes in their global dimensions. We also use molecular modelling and simulation techniques to uncover the mechanistic underpinnings of sequential phosphorylation of p53 TAD by kinase CK1delta. We identified a conserved, positively charged region on CK1delta that sequesters the negatively charged, phosphorylated p53 peptide, thereby constraining the positioning of the rest of the peptide for subsequent phosphorylation. Finally, we demonstrate that many critical scaffold proteins in phase-separated condensates are enriched in charge-absent regions. Altogether, this thesis contributes to a better understanding of how charge modulation by phosphorylation alters the structure, and thus function of IDPs.
|Date of Award
|1 Aug 2023
- The University of Manchester
|James Warwicker (Supervisor) & Perdita Barran (Supervisor)
- Intrinsically disordered proteins
- Molecular dynamics
- Computational chemistry