The Ca2+ Influence on Calmodulin Unfolding Pathway: A Steered Molecular Dynamics Simulation Study

Abstract

The force-induced unfolding of calmodulin (CaM) was investigated at atomistic details with steered molecular dynamics. The two isolated CaM domains as well as the full-length CaM were simulated in N-C-terminal pulling scheme, and the isolated N-lobe of CaM was studied specially in two other pulling schemes to test the effect of pulling direction and compare with relevant experiments. Both Ca2+-loaded CaM and Ca2+-free CaM were considered in order to define the Ca2+ influence to the CaM unfolding. The results reveal that the Ca2+ significantly affects the stability and unfolding behaviors of both the isolated CaM domains and the full-length CaM. In Ca2+-loaded CaM, N-terminal domain unfolds in priori to the C-terminal domain. But in Ca2+-free CaM, the unfolding order changes, and C-terminal domain unfolds first. The force-extension curves of CaM unfolding indicate that the major unfolding barrier comes from conquering the interaction of two EF-hand motifs in both N- and C- terminal domains. Our results provide the atomistic-level insights in the force-induced CaM unfolding and explain the observation in recent AFM experiments.

Characterization and identification of calmodulin and calmodulin binding proteins in hemocyte of the black tiger shrimp (Penaeus monodon)

“…This observation was similar to that of pearl oyster and freshwater mussel CaMs (Li et al, 2005;Ren et al, 2013). The binding of Ca 2+ ions could induce a conformational change of CaM to be compact, while apo-CaM may be in a less compact form, resulting in the different electrophoretic mobility (Zhang and Lou, 2012). This result was also supported by CD experiment, in that Ca 2+ ions increased α-helical content of rPmCaM when compared to that of the apoprotein.…”

Crystal structure of Arabidopsis thaliana calmodulin7 and insight into its mode of DNA binding

“…We therefore expect that removal of Ca 2+ from EF3 or EF-1 would disrupt the structural integrity of the CaM, hence DNA binding [17]. Moreover, previous molecular dynamics simulation indicated that removal of even a single Ca 2+ ion could induce unfolding of CaM, and hence the overall structure adopts a closed conformation [2]. Therefore, we expect that mutation of even a single EF-hand motif is sufficient to change the conformation of CaM, and when done in the C-terminal domain would abolish structural integrity of the C-terminal domain and thus the DNA interactions [17].…”

“…CaM plays essential role in Ca 2+ signaling, regulating numerous intracellular processes such as cell motility, growth, proliferation, and apoptosis . CaM binds Ca 2+ ions using its helix‐loop‐helix (EF‐hand) structural motif, and this motif generally undergoes large conformational changes upon Ca 2+ binding . In the Ca 2+ ‐loaded form, the CaM adopts stable state, and each EF‐hand opens so that its two alpha helices become perpendicular to each other.…”

Comparison of ligand binding and conformational stability of human calmodulin with its homolog from the malaria parasitePlasmodium falciparum

“…Several studies proposed higher stability for the N-domain in both apo-and CaCaM attributing the transitions with higher heat capacities to the N-lobe. 4,5,32 In contrast, other authors suggested a reverse order of stability in apo-and CaCaM 2,3 F I G U R E 5 Heat stability of human and Pf CaM. Thermal stability was studied by DSC in standard assay buffer.…”

“…The C-terminal domain bears higher affinity Ca 2+ -binding sites, so it is expected to possess higher stability in Ca 2+saturated CaM (CaCaM) as it was proposed in studies using experimental or computational approaches. 2,3 In contrast, higher stability for the N-terminal domain (N-domain) in both apo and holo CaM forms was suggested based on thermal melting profiles observed with differential scanning calorimetry. 4,5 Ca 2+ -binding to CaM results in a global conformational change involving rearrangement of the helices as well as additional helix formation.…”