Electron Cryo-Microscopic Study of the Type 1 IP3R

Inositol 1,4,5-trisphosphate receptors (IP3Rs) play important roles in a battery of cellular activities. Structural study of the receptors is therefore very important for understanding how they are gated by their natural ligands and are modulated by their intracellular partners. In past several years, multiple groups have generated very disparate reconstructions of the type 1 IP3R since the first low-resolution cryoEM structure was published (Jiang, et al EMBO J. 2002). Striking structural variations have been reported for receptors in different detergents and for receptors prepared from native and sf9 cells by the same research groups, suggesting that the biochemical preparations of the receptors have significant variations and that the heterogeneity in the samples could be a limiting factor in reaching accordant results. To help resolve such discrepancies, we are conducting single particle reconstruction of the type 1 IP3Rs from both native tissues and heterogeneous expression systems. We worked out conditions that maintained the stability and biochemical homogeneity of the purified receptors, and introduced site-specific mutations and prepared accessory binding partners to facilitate the mapping of specific locations in the 3D reconstruction. We are currently collecting datasets from the receptors in different conditions in order to examine the consistency among the reconstructed structures.