Structural basis for preferential Ca²⁺-displacement by a paramagnetic ion (Ce³⁺) in non-myristoylated neuronal calcium sensor-1

Abstract

We have investigated the Ca²⁺-displacement by a trivalent lanthanide ion (Ce³⁺) in a Ca²⁺-sensor protein called neuronal calcium sensor-1 in its non-myristoylated form by NMR. We observe that all the Ca²⁺ bound to the three active Ca²⁺-binding loops are displaced by Ce³⁺. Microscopically, NMR paramagnetic titration data on [Ca²⁺]-non-myr-NCS-1 shows that the Ce³⁺ displaces Ca²⁺ first from both sites, EF2 and EF3, almost in parallel, followed by the displacement from EF4. Between EF2 and EF3, the NMR data conclusively shows that the Ca²⁺-displacement is more prominent in EF2 as compared to that in EF3. These findings have been attributed to: (i) the presence of a higher number of negatively charged residues in EF2 compared to EF3 and EF4, and, (ii) the presence of a Lys instead of a highly conserved Phe at –4 position in the first Ca²⁺-binding loop, which results in a relatively more open conformation of the N-terminal domain compared to the C-terminal counterpart.