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Title: Biophysical investigations of PKD domains : mechanosensation and pathogenesis
Author: Forman, J. R.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2006
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Autosomal dominant polycystic kidney disease is one of the most common human genetic disorders, and most cases are linked to mutations in the protein polycystin-1. In 2003, polycystin-1 was proposed to function as a mechanosensor, sensing changes in kidney tubule fluid flow. This dissertation presents investigations into the mechanical properties of PKD domains, which constitute approximately half of the extracellular region of the transmembrane polycystin-1 protein. Using single molecule force spectroscopy, PKD domains of polycystin-1 were shown to exhibit a phenomenal resistance to unfolding under applied external forces. These results demonstrate that PKD domains would remain folded under the physiological forces of fluid flow and suggest that polycystin-1 has evolved to fulfil a mechanical function. How do proteins resist unfolding under applied forces? Computer simulations, protein engineering, and force spectroscopy were employed to investigate the unfolding pathway of PKD domains under force. This work reveals the formation of force-induced non-native contacts in the PKD domains. These non-native contacts appear to be the critical feature for stabilising PKD domains against unfolding. Finally, this dissertation presents the thermodynamic effects of pathogenic mutations in the PKD domains of polycystin-1. These data suggest that a significant proportion of pathogenic mutations will act to destabilise protein domains, sometimes preventing folding. Thermodynamic destabilisation is likely to be the molecular pathogenic mechanism for a large fraction of genetic diseases. Further research in this vein, to elucidate the molecular biophysical effects of mutations, will be vital for our understanding of genetic diseases.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available