Inositol 1,4,5-trisphosphate-gated calcium channel ITPR3
calcium ion binding
Structural titration reveals Ca<sup>2+</sup>-dependent conformational landscape of the IP<sub>3</sub> receptor.
Inositol 1,4,5-trisphosphate receptors (IP<sub>3</sub>Rs) are endoplasmic reticulum Ca<sup>2+</sup> channels whose biphasic dependence on cytosolic Ca<sup>2+</sup> gives rise to Ca<sup>2+</sup> oscillations that regulate fertilization, cell division and cell death. Despite the critical roles of IP<sub>3</sub>R-mediated Ca<sup>2+</sup> responses, the structural underpinnings of the biphasic Ca<sup>2+</sup> dependence that underlies Ca<sup>2+</sup> oscillations are incompletely understood. Here, we collect cryo-EM images of an IP<sub>3</sub>R with Ca<sup>2+</sup> concentrations spanning five orders of magnitude. Unbiased image analysis reveals that Ca<sup>2+</sup> binding does not explicitly induce conformational changes but rather biases a complex conformational landscape consisting of resting, preactivated, activated, and inhibited states. Using particle counts as a proxy for relative conformational free energy, we demonstrate that Ca<sup>2+</sup> binding at a high-affinity site allows IP<sub>3</sub>Rs to activate by escaping a low-energy resting state through an ensemble of preactivated states. At high Ca<sup>2+</sup> concentrations, IP<sub>3</sub>Rs preferentially enter an inhibited state stabilized by a second, low-affinity Ca<sup>2+</sup> binding site. Together, these studies provide a mechanistic basis for the biphasic Ca<sup>2+</sup>-dependence of IP<sub>3</sub>R channel activity.