Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket

Overview

Journal Int J Mol Sci

Publisher MDPI

Specialties Biochemistry
Chemistry
Molecular Biology

Date 2023 Nov 14

PMID 37958614

Authors

Angelos Thanassoulas

Maria Theodoridou

Laila Barrak

Emna Riguene

Tamader Alyaarabi

Mohamed A Elrayess

F Anthony Lai

Michail Nomikos

Affiliations

Soon will be listed here.

Abstract

Calmodulin (CaM) is a small, multifunctional calcium (Ca)-binding sensor that binds and regulates the open probability of cardiac ryanodine receptor 2 (RyR2) at both low and high cytosolic Ca concentrations. Recent isothermal titration calorimetry (ITC) studies of a number of peptides that correspond to different regions of human RyR2 showed that two regions of human RyR2 (3584-3602aa and 4255-4271aa) bind with high affinity to CaM, suggesting that these two regions might contribute to a putative RyR2 intra-subunit CaM-binding pocket. Moreover, a previously characterized de novo long QT syndrome (LQTS)-associated missense CaM mutation (E105A) which was identified in a 6-year-old boy, who experienced an aborted first episode of cardiac arrest revealed that this mutation dysregulates normal cardiac function in zebrafish by a complex mechanism that involves alterations in both CaM-Ca and CaM-RyR2 interactions. Herein, to gain further insight into how the CaM E105A mutation leads to severe cardiac arrhythmia, we generated large quantities of recombinant CaM and CaM proteins. We then performed ITC experiments to investigate and compare the interactions of CaM and CaM mutant protein with two synthetic peptides that correspond to the two aforementioned human RyR2 regions, which we have proposed to contribute to the RyR2 CaM-binding pocket. Our data reveal that the E105A mutation has a significant negative effect on the interaction of CaM with both RyR2 regions in the presence and absence of Ca, highlighting the potential contribution of these two human RyR2 regions to an RyR2 CaM-binding pocket, which may be essential for physiological CaM/RyR2 association and thus channel regulation.

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