A, Cartoon depicting CaM interaction domains on the CaV1.2 calcium channel (CI segment). Below, construct schematics depicting FRET interaction pairs used for binding assessment.
B, The canonical FRET binding curve between the CI region and CaMWT. In particular, an acceptor-centric FRET efficiency (EA) is plotted as a function of the relative free concentration of donor-tagged molecules Dfree (cerulean-CaMWT). Kd/EFF for CaMWT binding to CI region is 12000 Dfree units [5].
C-E, FRET binding curves between the CI region and CaMD96V (C), CaMD130G (D) and CaMF142L (E). Kd/EFF are 4000, 6500, and 1000 Dfree units, respectively. The binding curve for CaMWT is reproduced in gray for reference.