Construction and molecular dynamics simulation of calmodulin in the extended and in a bent conformation

Abstract

Analysis of sequence similarity and comparison of the three-dimensional (3D) structures of troponin C and calmodulin have revealed a sequence in the central helix of calmodulin with a high probability for bending. The three amino acids known to form a bend in the N-terminal portion of troponin C are also found in the central helix of calmodulin. The modelling of a bent calmodulin structure, using the dihedral angles of the three residues in the bend of troponin C as a 3D template, results in a conformation of calmodulin where the N-and C-terminal domains are able to form contacts. Dynamics simulations starting from the X-ray structure of calmodulin and from the modelled bent calmodulin were carried out to compare flexibility and correlated movements of CaZ+ in the binding loops. Both conformations of calmodulin remained stable during the period of simulation. In the simulation of calmodulin in the extended form, the motions of the Ca2+ atoms in the two domains (Ca2'l and CaZ ' 2 in one domain, and Ca2+3 and Ca2+4 in the other) are correlated. In the simulation of the bent form, an additional correlation between the Ca atoms in the two different domains is observed. The results are compatible with the occurrence of a bent conformation of calmodulin in the presence of targets, and with increased Ca2+ affinity and cooperativity of the CaZ + -binding loops in the calmodulin-peptide complexes.

Oligomerization of the plasma membrane calcium pump involves two regions with different thermal stability

“…This result indicates that: (a) the region of the pump involved in the dimerization and preserved after 7 h of denaturation at 44³C does not correspond to the CaM-binding domain and; (b) the association only occurs if at least the CaM-binding domain of one of the monomers has its native structure preserved. This last statement was con¢rmed by the progressive loss of energy transfer after the addition of increasing CaM concentrations to the native enzyme (Table 2), agreeing with Vorherr et al [8] who showed that CaM tends to maintain the enzyme in the monomeric state hindering its oligomerization.…”

“…On the other hand, it was demonstrated that the dimeric calcium pump is able to bind one CaM molecule [8]. In that paper, it was suggested that this result could be compatible with either the binding of the CaM molecule to only one monomer of the dimeric pump, or with its simultaneous binding to both monomers.…”

“…Structural studies on the synthetic peptide C28W, which sequence corresponds to the above mentioned domain, showed that this fragment has a strong tendency to form aggregates and, under special conditions, to form dimers [9]. By analogy, it was proposed that this interaction could be responsible for the dimerization [8]. However, there is no experimental evidence supporting this hypothesis.…”

“…This result indicates that the structure of a protein region involved in the oligomerization is preserved after 7 h of thermal denaturation at 44³C, but it is damaged when heating under stronger conditions. It has been proposed that the CaM-binding domain mediates the self-association of the PMCA [8]. According to the results shown in Fig.…”

“…Vorherr et al [8] demonstrated that the calmodulin (CaM)-binding domain of the pump is involved in the oligomerization process. Structural studies on the synthetic peptide C28W, which sequence corresponds to the above mentioned domain, showed that this fragment has a strong tendency to form aggregates and, under special conditions, to form dimers [9].…”