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Investigating the Pathogenesis of MYH7 Mutation Gly823Glu in Familial Hypertrophic Cardiomyopathy using a Mouse Model
Published on: August 8, 2022
Biomolecular interactions between human recombinant beta-MyHC and cMyBP-Cs implicated in familial hypertrophic
Jeanne Flavigny1, Philippe Robert, Jean-Claude Camelin
1INSERM U582, Institut de Myologie, Bâtiment Babinski, CHU Pitié-Salpêtrière, 47 Bld de l'Hôpital, 75651 Paris Cedex 13, France.
Insights
Familial hypertrophic cardiomyopathy (FHC) mutations can truncate cardiac myosin-binding protein C (cMyBP-C). Truncated cMyBP-C’s interaction with beta-MyHC depends on the truncation size, impacting disease development.
Area of Science:
- Biochemistry
- Molecular Biology
- Cardiovascular Research
Background:
- Cardiac myosin-binding protein C (cMyBP-C) is a key sarcomeric protein.
- Mutations in cMyBP-C are linked to familial hypertrophic cardiomyopathy (FHC).
- Many FHC mutations result in truncated cMyBP-C variants.
Purpose of the Study:
- To investigate the in vitro interaction between FHC-associated truncated cMyBP-C mutants and human beta-myosin heavy chain (beta-MyHC).
- To determine if the extent of cMyBP-C truncation affects its binding to beta-MyHC.
Main Methods:
- Production of recombinant wild-type and truncated cMyBP-C proteins using a baculovirus/insect cell system.
- Purification of proteins via metal affinity chromatography.
- Real-time analysis of protein interactions using biosensor technology with immobilized anti-beta-MyHC antibodies.
Main Results:
- Wild-type cMyBP-C and a mutant lacking half of the C10 domain interacted with beta-MyHC.
- Two mutants with truncations in the C5-C9 region showed no detectable interaction with beta-MyHC.
- The size of the C-terminal truncation significantly influences cMyBP-C binding to beta-MyHC.
Conclusions:
- Biosensor technology effectively analyzes cMyBP-C and beta-MyHC interactions in vitro.
- The study demonstrates that the degree of cMyBP-C truncation is critical for its interaction with beta-MyHC.
- Impaired interaction due to truncation may lead to protein degradation and contribute to FHC pathogenesis.
Objective:
Cardiac myosin-binding protein C (cMyBP-C) is a component of sarcomere that contains at least three putative myosin-binding sites. Mutations in its gene are implicated in familial hypertrophic cardiomyopathy (FHC) and most of them are predicted to produce C-terminal truncated cMyBP-Cs. The aim of the present study was to analyze whether cMyBP-C truncated mutants resulting from FHC mutations interact in vitro with human beta-MyHC.
Methods:
Recombinant proteins were produced using the baculovirus/insect cell system, and wild type and three truncated cMyBP-Cs were purified using metal affinity chromatography. The interaction between recombinant proteins was analyzed in real time using biosensor technology on immobilized anti-beta-MyHC antibodies.
Results:
Biomolecular interaction with beta-MyHC was detected for both wild type cMyBP-C and a truncated mutant lacking half of the C-terminal C10 domain. In contrast, no interaction with beta-MyHC was found for two truncated cMyBP-Cs lacking at least the C5-C9 region.
Conclusions:
Biosensor technology allows in vitro analysis of the interaction between human beta-MyHC and cMyBP-C mutants resulting from FHC mutations. The data show that the interaction depends on the size of the truncation. This suggests that, in the context of FHC, impairment of suitable interaction between beta-MyHC and some of the truncated cMyBP-Cs may promote degradation of the truncated proteins and therefore contribute to the development of the disease.
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