Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket
Calmodulin (CaM) is a small, multifunctional calcium (Ca<sup>2+</sup>)-binding sensor that binds and regulates the open probability of cardiac ryanodine receptor 2 (RyR2) at both low and high cytosolic Ca<sup>2+</sup> concentrations. Recent isothermal titration calorimetry (I...
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MDPI AG
2023-10-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/21/15630 |
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| author | Angelos Thanassoulas Maria Theodoridou Laila Barrak Emna Riguene Tamader Alyaarabi Mohamed A. Elrayess F. Anthony Lai Michail Nomikos |
| author_facet | Angelos Thanassoulas Maria Theodoridou Laila Barrak Emna Riguene Tamader Alyaarabi Mohamed A. Elrayess F. Anthony Lai Michail Nomikos |
| author_sort | Angelos Thanassoulas |
| collection | DOAJ |
| description | Calmodulin (CaM) is a small, multifunctional calcium (Ca<sup>2+</sup>)-binding sensor that binds and regulates the open probability of cardiac ryanodine receptor 2 (RyR2) at both low and high cytosolic Ca<sup>2+</sup> 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<sup>2+</sup> 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<sup>WT</sup> and CaM<sup>E105A</sup> proteins. We then performed ITC experiments to investigate and compare the interactions of CaM<sup>WT</sup> and CaM<sup>E105A</sup> 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<sup>2+</sup>, 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. |
| first_indexed | 2024-03-11T11:29:05Z |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-11T11:29:05Z |
| publishDate | 2023-10-01 |
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| spelling | doaj.art-c41c817fd739420fa0534953dca00f342023-11-10T15:04:41ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672023-10-0124211563010.3390/ijms242115630Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding PocketAngelos Thanassoulas0Maria Theodoridou1Laila Barrak2Emna Riguene3Tamader Alyaarabi4Mohamed A. Elrayess5F. Anthony Lai6Michail Nomikos7College of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarBiomedical Research Center, Qatar University, Doha P.O. Box 2713, QatarCollege of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarCollege of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarCollege of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarCollege of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarCollege of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarCollege of Medicine, QU Health, Qatar University, Doha P.O. Box 2713, QatarCalmodulin (CaM) is a small, multifunctional calcium (Ca<sup>2+</sup>)-binding sensor that binds and regulates the open probability of cardiac ryanodine receptor 2 (RyR2) at both low and high cytosolic Ca<sup>2+</sup> 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<sup>2+</sup> 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<sup>WT</sup> and CaM<sup>E105A</sup> proteins. We then performed ITC experiments to investigate and compare the interactions of CaM<sup>WT</sup> and CaM<sup>E105A</sup> 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<sup>2+</sup>, 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.https://www.mdpi.com/1422-0067/24/21/15630calmodulinryanodine receptorRyR2arrhythmiascardiac disease |
| spellingShingle | Angelos Thanassoulas Maria Theodoridou Laila Barrak Emna Riguene Tamader Alyaarabi Mohamed A. Elrayess F. Anthony Lai Michail Nomikos Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket International Journal of Molecular Sciences calmodulin ryanodine receptor RyR2 arrhythmias cardiac disease |
| title | Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket |
| title_full | Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket |
| title_fullStr | Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket |
| title_full_unstemmed | Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket |
| title_short | Arrhythmia-Associated Calmodulin E105A Mutation Alters the Binding Affinity of CaM to a Ryanodine Receptor 2 CaM-Binding Pocket |
| title_sort | arrhythmia associated calmodulin e105a mutation alters the binding affinity of cam to a ryanodine receptor 2 cam binding pocket |
| topic | calmodulin ryanodine receptor RyR2 arrhythmias cardiac disease |
| url | https://www.mdpi.com/1422-0067/24/21/15630 |
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