Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.757813
Title: Small heat shock protein interactions with in vivo partners
Author: Collier, Miranda
ISNI:       0000 0004 7430 6221
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2018
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Abstract:
Small heat-shock proteins (sHsps) are part of a broad cellular sys- tem that functions to maintain a stable proteome under stress. They also perform a variety of regulatory roles at physiological conditions. Despite the multitude of sHsp targets, their interactions with partners are not well understood due to highly dynamical structures. In this thesis, I apply a variety of biophysical and structural approaches to examine distinct interactions made by the abundant human sHsps αβ-crystallin and Hsp27. First, I find that αβ-crystallin binds a cardiac-specific domain of the muscle sarcomere protein titin. A cardiomyopathy-causative variant of αβ-crystallin is shown to disrupt this interaction, with demonstrated implications for tissue biomechanics. Next, I investigate the conformation and unfolding behaviour of another sarcomere-associated protein, filamin C, finding support for the hypothesis that it is mechanosensitive. This leads into an interrogation of the interaction between filamin C and Hsp27, which we find is modulated by phosphorylation of Hsp27. This modulation only manifests during filamin C unfolding, pointing toward a protective chaperoning mode against over-extension during mechanical stress. This finding is bolstered by up-regulation and interaction of both proteins in a mouse model of heart failure. I establish a system for similar studies of a third sHsp, cvHsp, which is muscle-specific and implicated in various myopathies but scantly understood at the molecular level compared to αβ-crystallin and Hsp27. Finally, I probe the stoichiometries and kinetics of complexes formed between αβ-crystallin and Hsp27 themselves, which co-assemble into a highly polydisperse ensemble. This involved the development of a high-resolution native mass spectrometry method for disentangling heterogeneous systems. Together these findings add to our understanding of the roles and mechanisms of ATP-independent molecular chaperones.
Supervisor: Benesch, Justin Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.757813  DOI: Not available
Keywords: Biophysics--Research ; Cardiovascular disease ; Native mass spectrometry ; Molecular chaperones ; Mechanobiology ; Heat shock proteins
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