Investigation of non-covalent complexations of Ca(II) and Mg(II) ions with insulin by using electrospray ionization mass spectrometry.

Rationale: Insulin is a peptide hormone secreted by pancreatic β-cells. Ca(II) and Mg(II) ions play an important role in the secretion of insulin. There is no study about a direct complexation of Ca(II) or Mg(II) with insulin and their equilibrium constants. Electrospray ionization mass spectrometry (ESI-MS) is a practical method for the monitoring of non-covalent complexes such as Ca(II)-insulin and Mg(II)-insulin. Here, the equilibrium constants of Ca(II)-insulin and Mg(II)-insulin non-covalent complexes have been calculated after ESI-MS measurements in aqueous solutions.

Methods: The effects of pH, competitive binding, ion exchange, and Na(I) and K(I) ions on Ca(II)-insulin and Mg(II)-insulin complexation have been examined by measuring by ESI-MS. The dissociation equilibrium constants (K1 and K2 ) of Ca(II)-insulin and Mg(II)-insulin complexes were calculated from the binomial graph derived from the ESI-MS normalized peak intensities. The MS/MS spectra of the complexes have been examined.

Results: The dissociation equilibrium constants were found to K1 : 1.29 × 10(-4)  M and K2 : 9.69 × 10(-4)  M for the Ca(II)-insulin complexes, and K1 : 1.37 × 10(-4)  M and K2 : 9.12 × 10(-4)  M for Mg(II)-insulin complexes. Ca(II) ions have higher complexation capability with insulin than Mg(II) ions.

Conclusions: The binding equilibrium constants of Ca(II)- and Mg(II)-insulin non-covalent complexes have been determined successfully by ESI-MS. Ca(II) and Mg(II) ions are involved in the insulin secretion by forming non-covalent complexes. Copyright © 2016 John Wiley & Sons, Ltd.