Calcium binding characteristics and structural changes of phosvitin

• Phosvitin exhibited distinctly high and low affinity calcium-binding modes.
• The high affinity binding sites contained nearly 30 mol calcium/mol phosvitin.
• The low affinity binding mode exhibited many calcium was sequestered by phosvitin.
• Conformation changes of phosvitin were opposite in these two types of binding modes.

Phosvitin is a unique highly phosphorylated protein that plays a role in the regulation of calcification. We conducted a comprehensive study of the chemical, thermodynamic and structural aspects of the interaction of phosvitin with calcium ions using a calcium ion selective electrode (ISE), isothermal titration calorimetry (ITC), circular dichroism spectrum (CD) and fluorescence spectroscopy, respectively. The results showed that under neutral and alkaline conditions, distinct high affinity and low affinity binding modes existed in the interaction between phosvitin and calcium. The high affinity association constant was approximately 104 mol− 1, while the binding sites contained nearly 30 mol of calcium per mole of phosvitin. This reaction was driven by enthalpy. The unordered and β-turn conformations of phosvitin increased, while the β-sheet conformation decreased. The main interaction forces were electrostatic force, hydrogen bonds or van der Waals force. The low affinity association constant and binding sites were not constant, as many calcium ions were sequestered by phosvitin. The binding reaction was driven by entropy, and the β-sheet conformation of phosvitin increased while the unordered conformation decreased. The main interaction force was hydrophobic force. However, under acidic conditions, the interaction between phosvitin and calcium was an entropy-driven endothermic reaction, and the main interaction force was weak hydrophobic force. This calcium-binding characteristic of phosvitin may play a specific role in its biological function.

Here we conducted a comprehensive study of the chemical, thermodynamic and structural aspects of the interaction of phosvitin with calcium ions. Distinct high affinity and low affinity binding modes existed in the phosvitin-Ca2+ interaction, driving by enthalpy and entropy, respectively. In addition, the conformation of phosvitin exhibited opposite changes.Figure optionsDownload as PowerPoint slide