The Lqt-Associated Calmodulin Mutant E141g Induces Disturbed Ca2+-Dependent Binding And A Flickering Gating Mode Of The Ca(V)1.2 Channel

Calmodulin (CaM) mutations are associated with congenital long QT (LQT) syndrome (LQTS), which may be related to the dysregulation of the cardiac-predominant Ca2+ channel isoform Ca(V)1.2. Among various mutants, CaM-E141G was identified as a critical missense variant. However, the interaction of this CaM mutant with the Ca(V)1.2 channel has not been determined. In this study, by utilizing a semiquantitative pull-down assay, we explored the interaction of CaM-E141G with CaM-binding peptide fragments of the Ca(V)1.2 channel. Using the patch-clamp technique, we also investigated the electrophysiological effects of the mutant on Ca(V)1.2 channel activity. We found that the maximum binding (B-max) of CaM-E141G to the proximal COOH-terminal region, PreIQ-IQ, PreIQ, IQ, and NT (an NH2-terminal peptide) was decreased (by 17.71-59.26%) compared with that of wild-type CaM (CaM-WT). In particular, the Ca2+-dependent increase in B-max became slower with the combination of CaM-E141G + PreIQ and IQ but faster in the case of NT. Functionally, CaM-WT and CaM-E141G at 500 nM Ca2+ decreased Ca(V)1.2 channel activity to 24.88% and 55.99%, respectively, compared with 100 nM Ca2+, showing that the inhibitory effect was attenuated in CaM-E141G. The mean open time of the Ca(V)1.2 channel was increased, and the number of blank traces with no channel opening was significantly decreased. Overall, CaM-E141G exhibits disrupted binding with the Ca(V)1.2 channel and induces a flickering gating mode, which may result in the dysfunction of the Ca(V)1.2 channel and, thus, the development of LQTS. The present study is the first to investigate the detailed binding properties and single-channel gating mode induced by the interaction of CaM-E141G with the Ca(V)1.2 channel.