Exploring the heart of MyBP-C

Student thesis: Doctoral ThesisDoctor of Philosophy


Mutations in the sarcomeric protein cardiac myosin binding protein C (cMyBP-C) are one of the most common causes of hereditary hypertrophic cardiomyopathy (HCM). cMyBP-C is made up of 11 immunoglobulin-like or fibronectin-like domains, designated C0-C10, and has been found to interact with myosin, actin and titin. The N-terminus contains binding sites for both actin and myosin, suggesting it plays a role in the regulation of actomyosin cross-bridge cycling kinetics, which is the major role identified for MyBP-C. The C-terminus of the protein is thought to target MyBP-C to the thick filament backbone due to the fact that it contains binding sites for both myosin and titin. The function of the central domains (C3-C6) has not yet been elucidated; however the largest number of HCM-linked mutations are found in the C6 domain, followed by the C5 domain, intimating that this region does have an important functional role. The aims of this project were therefore:

1. To elucidate the structure and functional role of the central domains of cMyBP-C.
2. To understand the mechanism of disease of HCM-linked missense mutations in the C3C6 region.

To gain a better understanding of the role of the central domains a yeast two hybrid (Y2H) assay was carried out to identify putative novel binding partners. Recombinantly expressed fragments of the central domains were also used in biochemical assays in order to determine whether this region could be involved in interactions with actin. Constructs containing HCM-linked missense mutations within the central domain region were also generated and western blot analysis and immunofluorescence microscopy was utilised in order to gain insights into possible mechanisms of disease.

Whilst the results of these studies identified no novel binding partners via the Y2H screen, there was some evidence of an interaction between the central domains and actin. Furthermore, biochemical assays suggested that the central region could have a role in the regulation of actomyosin cross-bridge cycling kinetics. Experiments utilising HCM-linked missense mutation containing constructs highlighted the fact that there seems to be no unified disease mechanism by which these mutations lead to HCM. Instead it appears that these mutations lead to HCM via different pathways.

Further study of a wider range of missense mutations should therefore be carried out in order to inform on the function of specific regions of the protein, as well as allow for classification of different mutations depending on their pathogenesis to create more personalised methods of treatment.
Date of Award1 Dec 2021
Original languageEnglish
Awarding Institution
  • King's College London
SupervisorElisabeth Ehler (Supervisor) & Thomas Kampourakis (Supervisor)

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