Apo and InsP3-bound crystal structures of the ligand-binding domain of an InsP3 receptor

Department of Physiology, Howard Hughes Medical Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Nature Structural & Molecular Biology (Impact Factor: 13.31). 09/2011; 18(10):1172-4. DOI: 10.1038/nsmb.2112
Source: PubMed


We report the crystal structures of the ligand-binding domain (LBD) of a rat inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R) in its apo and InsP3-bound conformations. Comparison of these two conformations reveals that LBD's first β-trefoil fold (β-TF1) and armadillo repeat fold (ARF) move together as a unit relative to its second β-trefoil fold (β-TF2). Whereas apo-LBD may spontaneously transition between gating conformations, InsP3 binding shifts this equilibrium towards the active state.

Download full-text


Available from: Kyuwon Baek, Apr 27, 2014
  • Source
    • "and B) [15] [16]. Other evidence for interactions between the ligand-binding domains and IP 3 -induced conformational changes comes from NMR and small-angle X-ray scattering studies performed when these domains are expressed as independent entities [55]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The ability of cells to maintain low levels of Ca(2+) under resting conditions and to create rapid and transient increases in Ca(2+) upon stimulation is a fundamental property of cellular Ca(2+) signaling mechanism. An increase of cytosolic Ca(2+) level in response to diverse stimuli is largely accounted for by the inositol 1,4,5-trisphosphate receptor (IP3R) present in the endoplasmic reticulum membranes of virtually all eukaryotic cells. Extensive information is currently available on the function of IP3Rs and their interaction with modulators. Very little, however, is known about their molecular architecture and therefore most critical issues surrounding gating of IP3R channels are still ambiguous, including the central question of how opening of the IP3R pore is initiated by IP3 and Ca(2+). Membrane proteins such as IP3R channels have proven to be exceptionally difficult targets for structural analysis due to their large size, their location in the membrane environment, and their dynamic nature. To date, a 3D structure of complete IP3R channel is determined by single-particle cryo-EM at intermediate resolution, and the best crystal structures of IP3R are limited to a soluble portion of the cytoplasmic region representing ∼15% of the entire channel protein. Together these efforts provide the important structural information for this class of ion channels and serve as the basis for further studies aiming at understanding of the IP3R function.
    Full-text · Article · Sep 2014 · Cell Calcium
  • Source
    • "As with all ion channels, activation of IP3R proceeds via re-arrangements of interactions between protein domains within the oligomeric channel. For IP3R, these conformational changes are initiated by IP3 binding to the IP3-binding core (IBC, residues 224–604) of each of the four IP3R subunits, and then proceed via re-organization of intramolecular interactions between the IBC and suppressor domain (SD, residues 1–223) [12], [13]. These conformational changes are proposed to disrupt contacts between the N-terminal regions of the four subunits and to culminate in opening of the Ca2+-permeable pore [13]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Interactions between proteins are a hallmark of all cellular activities. Such interactions often occur with low affinity, a feature that allows them to be rapidly reversible, but it makes them difficult to detect using conventional methods such as yeast 2-hybrid analyses, co-immunoprecipitation or analytical ultracentrifugation. We developed a simple and economical bead aggregation assay to study low-affinity interactions between proteins. By coating beads with interacting proteins, the weak interactions between many proteins are sufficient to allow stable aggregation of beads, an avidity effect. The aggregation is easily measured to allow quantification of protein-protein interactions under a variety of controlled conditions. We use this assay to demonstrate low-affinity interactions between the N-terminal domains of an intracellular Ca(2+) channel, the type 1 inositol 1,4,5-trisphosphate receptor. This simple bead aggregation assay may have widespread application in the study of low-affinity interactions between macromolecules.
    Full-text · Article · Mar 2013 · PLoS ONE
  • Source
    • "One possibility is that an endogenous EF-hand-like structure might provide the Ca2+-binding site and that its interaction with the 1-8-14 motif links IP3 and Ca2+ binding (Figure 7A). Bioinformatic analyses had suggested the presence of two possible EF-hand-like structures within the IBC [9,54], but neither is evident in high-resolution structures of the IBC [55] and NT [5,56]. Neither have we succeeded in identifying a complementary partner of the 1-8-14 motif. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Binding of inositol 1,4,5-trisphosphate (IP3) to the IP3-binding core (residues 224-604) of IP3 receptors (IP3R) initiates opening of these ubiquitous intracellular Ca2+ channels. The mechanisms are unresolved but require conformational changes to pass through the suppressor domain (residues 1-223). A calmodulin-binding peptide derived from myosin light chain kinase uncouples these events. We identified a similar conserved 1-8-14 calmodulin-binding motif within the suppressor domain of IP3R1 and using peptides and mutagenesis we demonstrate that it is essential for IP3R activation, whether assessed by IP3-evoked Ca2+ release or patch-clamp recoding of nuclear IP3R. Mimetic peptides specifically inhibit activation of IP3R by uncoupling the IP3-binding core from the suppressor domain. Mutations of key hydrophobic residues within the endogenous 1-8-14 motif mimic the peptides. Our results show that an endogenous 1-8-14 motif mediates conformational changes that are essential for IP3R activation. The inhibitory effects of calmodulin and related proteins may result from disruption of this essential interaction.
    Full-text · Article · Sep 2012 · Biochemical Journal
Show more