| Products/Services Used | Details | Operation |
|---|---|---|
| Peptide Synthesis | Isothermal titration calorimetry of RyR2(R3581-P3607) peptide with CaM - A peptide corresponding to the CaMBD (human RyR2 3581- RSKKAVWHKLLSKQRKRAVVACFRMAP-3607) was purchased from GenScript (Piscataway, USA) at > 95 % purity. Titration of the RyR2(R3581-L3607) peptide with CaMs was investigated under both Ca2+ free (apo) and Ca2+ saturating condition and followed using ITC. Purified CaM variants were dialyzed (10 K Slide-a-lyzerTM, Thermo Scientific) against the titration buffer (10 mM HEPES, 150 mM KCl, pH 7.2) with either 10 mM EDTA or 10 mM CaCl2 added for 40 h at 4 o C (with buffer changed at 24 h). | Get A Quote |
A number of point mutations in the intracellular Ca2+-sensing protein calmodulin (CaM) are arrhythmogenic, yet their underlying mechanisms are not clear. These mutations generally decrease Ca2+ binding to CaM and impair inhibition of CaMregulated Ca2+ channels like the cardiac Ca2+- release channel (ryanodine receptor, RyR2), and it appears that attenuated CaM Ca2+-binding correlates with impaired CaM-dependent RyR2 inhibition. Here, we investigated the RyR2 inhibitory action of the CaM p.Phe142Leu mutation (F142L; numbered including the start-Met), which markedly reduces CaM Ca2+-binding. Surprisingly, CaM-F142L had little to no aberrant effect on RyR2-mediated store-overload induced Ca2+- release in HEK293 ce... More
