3 years ago

Calmodulin mutant in central linker reduces the binding affinity with PreIQ and IQ while interacting with CaV1.2 channels

Yan Liu, Shan Yan, Sichong Chen, Jie Zhang, Yixuan Shen, Jingyang Su, Guilin He, Rui Feng, Dongxue Shao, Liying Hao

Calmodulin (CaM) was reported to interact with PreIQ and IQ of CaV1.2 channels, but to date, no explicit binding sites of CaM were illustrated. Therefore, in the present study, we firstly used MOE (Molecular Operating Environment) for protein-protein docking and we found that the most likely residues of CaM that play an important role in the interface are concentrated in central linker region. Next we examined the binding properties of CaM and its mutants to PreIQ and IQ by GST pull-down assays. Here we confirmed that CaM binds to PreIQ and IQ in a concentration-dependent and [Ca2+]-dependent manner. However, silencing the effect of N-lobe and C-lobe by mutating two Ca2+ binding sites of each lobe abolished [Ca2+]-dependence of CaM binding, but could not influence the combination. And the mutant in central linker reduced the binding of CaM/PreIQ and CaM/IQ especially at low [Ca2+]. We confirmed that N-lobe and C-lobe play vital role in sensing the change of Ca2+, and found that the central linker of CaM is involved in the binding of CaM to CaV1.2 channels in particular at low [Ca2+], not only participates in the combination with PreIQ, but also with IQ.

Publisher URL: https://www.sciencedirect.com/science/article/pii/S0006291X20304897

DOI: 10.1016/j.bbrc.2020.03.018

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