The Effect of Ca2+, Lobe-Specificity, and CaMKII on CaM Binding to NaV1.1

Int J Mol Sci. 2018 Aug 23;19(9):2495. doi: 10.3390/ijms19092495.

Abstract

Calmodulin (CaM) is well known as an activator of calcium/calmodulin-dependent protein kinase II (CaMKII). Voltage-gated sodium channels (VGSCs) are basic signaling molecules in excitable cells and are crucial molecular targets for nervous system agents. However, the way in which Ca2+/CaM/CaMKII cascade modulates NaV1.1 IQ (isoleucine and glutamine) domain of VGSCs remains obscure. In this study, the binding of CaM, its mutants at calcium binding sites (CaM12, CaM34, and CaM1234), and truncated proteins (N-lobe and C-lobe) to NaV1.1 IQ domain were detected by pull-down assay. Our data showed that the binding of Ca2+/CaM to the NaV1.1 IQ was concentration-dependent. ApoCaM (Ca2+-free form of calmodulin) bound to NaV1.1 IQ domain preferentially more than Ca2+/CaM. Additionally, the C-lobe of CaM was the predominant domain involved in apoCaM binding to NaV1.1 IQ domain. By contrast, the N-lobe of CaM was predominant in the binding of Ca2+/CaM to NaV1.1 IQ domain. Moreover, CaMKII-mediated phosphorylation increased the binding of Ca2+/CaM to NaV1.1 IQ domain due to one or several phosphorylation sites in T1909, S1918, and T1934 of NaV1.1 IQ domain. This study provides novel mechanisms for the modulation of NaV1.1 by the Ca2+/CaM/CaMKII axis. For the first time, we uncover the effect of Ca2+, lobe-specificity and CaMKII on CaM binding to NaV1.1.

Keywords: Ca2+; CaM; CaMKII; IQ domain; NaV1.1.

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Calcium / chemistry*
  • Calcium / metabolism
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2 / chemistry*
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2 / genetics
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2 / metabolism
  • Calmodulin / chemistry*
  • Calmodulin / genetics
  • Calmodulin / metabolism
  • Cloning, Molecular
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Genetic Vectors / chemistry
  • Genetic Vectors / metabolism
  • Glutathione Transferase / chemistry
  • Glutathione Transferase / genetics
  • Glutathione Transferase / metabolism
  • HEK293 Cells
  • Humans
  • Kinetics
  • Molecular Docking Simulation
  • Mutation
  • NAV1.1 Voltage-Gated Sodium Channel / chemistry*
  • NAV1.1 Voltage-Gated Sodium Channel / genetics
  • NAV1.1 Voltage-Gated Sodium Channel / metabolism
  • Phosphorylation
  • Protein Binding
  • Protein Conformation, alpha-Helical
  • Protein Conformation, beta-Strand
  • Protein Interaction Domains and Motifs
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Thermodynamics

Substances

  • Calmodulin
  • NAV1.1 Voltage-Gated Sodium Channel
  • Recombinant Fusion Proteins
  • SCN1A protein, human
  • Glutathione Transferase
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2
  • Calcium