Structural and dynamic insights into the subtype-specific IP3-binding mechanism of the IP3 receptor

There are three subtypes of vertebrate inositol 1,4,5-trisphosphate (IP₃) receptor (IP₃R), a Ca²⁺-release channel on the ER membrane - IP₃R1, IP₃R2, and IP₃R3 - each of which has a distinctive role in disease development. To determine the subtype-specific IP₃-binding mechanism, we compared the thermodynamics, thermal stability, and conformational dynamics between the N-terminal regions of IP₃R1 (IP₃R1-NT) and IP₃R3 (IP₃R3-NT) by performing circular dichroism (CD), isothermal titration calorimetry (ITC), and hydrogen-deuterium exchange mass spectrometry (HDX-MS). Previously determined crystal structures of IP₃R1-NT and HDX-MS results from this study revealed that both IP₃R1 and IP₃R3 adopt a similar IP₃-binding mechanism. However, several regions, including the α- and β-interfaces, of IP₃R1-NT and IP₃R3-NT show significantly different conformational dynamics upon IP₃ binding, which may explain the different IP₃-binding affinities between the subtypes. The importance of the interfaces for subtype-specific IP₃ binding is also supported by the different dynamic conformations of the two subtypes in the apo-states. Furthermore, IP₃R1-NT and IP₃R3-NT show different IP₃-binding affinities and thermal stabilities, but share similar thermodynamic properties for IP₃ binding. These results collectively provide new insights into the mechanism underlying IP₃ binding to IP₃Rs and the subtype-specific regulatory mechanism.