Target-peptide-induced changes in the structure and dynamics of calmodulin as probed by frequency domain fluorimetry of bound Tb(III).

Tb(III) luminescence is used to probe the conformational change induced in the calcium regulatory protein calmodulin upon binding to a target peptide. The luminescence lifetime for Tb(III) measured by frequency domain fluorimetry increases from 1278 microseconds for the calmodulin complex to 1496 microsecond for the complex of calmodulin and M13, a peptide derived from the calmodulin target protein myosin light chain kinase. The intensity of the Tb(III) emission increases over the solution value by a factor of 726 and 891 when bound to calmodulin and to the complex of calmodulin and M13 respectively. The sensitivity of the Tb(III) decay rate to deuterated solvent was also measured and is consistent with a single water molecule bound to the metal in both the calmodulin and calmodulin-M13 complex. The most dramatic change induced by M13 is the threefold reduction in the width of the Tb(III) lifetime distribution, which is interpreted to be a target-peptide-induced annealing of the previously flexible metal-binding site.