Binding of Ca2+ to calmodulin and its tryptic fragments: theory and experiment.

Abstract

The Ca(2+)-binding constant of the protein calmodulin is determined experimentally at different pH and varying salt concentration. By comparison to statistical mechanical simulation results, it is shown that the shift in binding constant upon addition of salt is almost quantitatively due to electrostatic interactions. Specific interactions as well as effects due to structural rearrangements of the protein seem to be less important, indicating a structurally rather conserved protein upon addition of salt and changes in pH. The tryptic fragments of calmodulin also bind calcium with high affinity, and the electrostatic effects seem likewise to dominate the binding constant shifts in these systems. It is also shown that the chemical potential of free calcium ions, which is normally dominated by the salt, is strongly influenced by the highly charged calmodulin molecule. This complicates a detailed comparison at low salt concentration, since it requires very accurate information on the salt and protein concentrations, which normally are not available.