skip to main content


The NSF Public Access Repository (NSF-PAR) system and access will be unavailable from 5:00 PM ET until 11:00 PM ET on Friday, June 21 due to maintenance. We apologize for the inconvenience.

This content will become publicly available on December 1, 2024

Title: Understanding the molecular mechanisms of odorant binding and activation of the human OR52 family

Structural and mechanistic studies on human odorant receptors (ORs), key in olfactory signaling, are challenging because of their low surface expression in heterologous cells. The recent structure of OR51E2 bound to propionate provided molecular insight into odorant recognition, but the lack of an inactive OR structure limited understanding of the activation mechanism of ORs upon odorant binding. Here, we determined the cryo-electron microscopy structures of consensus OR52 (OR52cs), a representative of the OR52 family, in the ligand-free (apo) and octanoate-bound states. The apo structure of OR52csreveals a large opening between transmembrane helices (TMs) 5 and 6. A comparison between the apo and active structures of OR52csdemonstrates the inward and outward movements of the extracellular and intracellular segments of TM6, respectively. These results, combined with molecular dynamics simulations and signaling assays, shed light on the molecular mechanisms of odorant binding and activation of the OR52 family.

more » « less
Award ID(s):
Author(s) / Creator(s):
; ; ; ; ; ; ; ; ; ; ; ;
Publisher / Repository:
Date Published:
Journal Name:
Nature Communications
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract

    Type A γ-aminobutyric acid receptors (GABAARs) are inhibitory pentameric ligand-gated ion channels in the brain. Many anesthetics and neurosteroids act through binding to the GABAAR transmembrane domain (TMD), but the structural basis of their actions is not well understood and no resting-state GABAAR structure has been determined. Here, we report crystal structures of apo and the neurosteroid anesthetic alphaxalone-bound desensitized chimeric α1GABAAR (ELIC-α1GABAAR). The chimera retains the functional and pharmacological properties of GABAARs, including potentiation, activation and desensitization by alphaxalone. The apo-state structure reveals an unconventional activation gate at the intracellular end of the pore. The desensitized structure illustrates molecular determinants for alphaxalone binding to an inter-subunit TMD site. These structures suggest a plausible signaling pathway from alphaxalone binding at the bottom of the TMD to the channel gate in the pore-lining TM2 through the TM1–TM2 linker. The study provides a framework to discover new GABAAR modulators with therapeutic potential.

    more » « less
  2. Abstract

    Structure and functions of S100 proteins are regulated by two distinct calcium binding EF hand motifs. In this work, we used solution‐state NMR spectroscopy to investigate the cooperativity between the two calcium binding sites and map the allosteric changes at the target binding site. To parse the contribution of the individual calcium binding events, variants of S100A12 were designed to selectively bind calcium to either the EF‐I (N63A) or EF‐II (E31A) loop, respectively. Detailed analysis of the backbone chemical shifts for wildtype protein and its mutants indicates that calcium binding to the canonical EF‐II loop is the principal trigger for the conformational switch between ‘closed’ apo to the ‘open’ Ca2+‐bound conformation of the protein. Elimination of binding in S100‐specific EF‐I loop has limited impact on the calcium binding affinity of the EF‐II loop and the concomitant structural rearrangement. In contrast, deletion of binding in the EF‐II loop significantly attenuates calcium affinity in the EF‐I loop and the structure adopts a ‘closed’ apo‐like conformation. Analysis of experimental amide nitrogen (15N) relaxation rates (R1,R2, and15N–{1H} NOE) and molecular dynamics (MD) simulations demonstrate that the calcium bound state is relatively floppy with pico–nanosecond motions induced in functionally relevant domains responsible for target recognition such as the hinge domain and the C‐terminal residues. Experimental relaxation studies combined with MD simulations show that while calcium binding in the EF‐I loop alone does not induce significant motions in the polypeptide chain, EF‐I regulates fluctuations in the polypeptide in the presence of bound calcium in the EF‐II loop. These results offer novel insights into the dynamic regulation of target recognition by calcium binding and unravels the role of cooperativity between the two calcium binding events in S100A12.

    more » « less
  3. CC chemokine receptor 2 (CCR2) is a part of the chemokine receptor family, an important class of therapeutic targets. These class A G-protein coupled receptors (GPCRs) are involved in mammalian signaling pathways and control cell migration toward endogenous CC chemokine ligands, named for the adjacent cysteine motif on their N terminus. Chemokine receptors and their associated ligands are involved in a wide range of diseases and thus have become important drug targets. CCR2, in particular, promotes the metastasis of cancer cells and is also implicated in autoimmunity-driven type-1 diabetes, diabetic nephropathy, multiple sclerosis, asthma, atherosclerosis, neuropathic pain, and rheumatoid arthritis. Although promising, CCR2 antagonists have been largely unsuccessful to date. Here, we investigate the effect of an orthosteric and an allosteric antagonist on CCR2 dynamics by coupling long-timescale molecular dynamics simulations with Markov-state model theory. We find that the antagonists shift CCR2 into several stable inactive conformations that are distinct from the crystal structure conformation and disrupt a continuous internal water and sodium ion pathway, preventing transitions to an active-like state. Several metastable conformations present a cryptic drug-binding pocket near the allosteric site that may be amenable to targeting with small molecules. Without antagonists, the apo dynamics reveal intermediate conformations along the activation pathway that provide insight into the basal dynamics of CCR2 and may also be useful for future drug design.

    more » « less
  4. Agashe, Deepa (Ed.)
    Abstract Independent origins of sociality in bees and ants are associated with independent expansions of particular odorant receptor (OR) gene subfamilies. In ants, one clade within the OR gene family, the 9-exon subfamily, has dramatically expanded. These receptors detect cuticular hydrocarbons (CHCs), key social signaling molecules in insects. It is unclear to what extent 9-exon OR subfamily expansion is associated with the independent evolution of sociality across Hymenoptera, warranting studies of taxa with independently derived social behavior. Here, we describe OR gene family evolution in the northern paper wasp, Polistes fuscatus, and compare it to four additional paper wasp species spanning ∼40 million years of evolutionary divergence. We find 200 putatively functional OR genes in P. fuscatus, matching predictions from neuroanatomy, and more than half of these are in the 9-exon subfamily. Most OR gene expansions are tandemly arrayed at orthologous loci in Polistes genomes, and microsynteny analysis shows species-specific gain and loss of 9-exon ORs within tandem arrays. There is evidence of episodic positive diversifying selection shaping ORs in expanded subfamilies. Values of omega (dN/dS) are higher among 9-exon ORs compared to other OR subfamilies. Within the Polistes OR gene tree, branches in the 9-exon OR clade experience relaxed negative (relaxed purifying) selection relative to other branches in the tree. Patterns of OR evolution within Polistes are consistent with 9-exon OR function in CHC perception by combinatorial coding, with both natural selection and neutral drift contributing to interspecies differences in gene copy number and sequence. 
    more » « less
  5. Abstract

    Molecular chaperones are ATP‐consuming machines, which facilitate the folding of proteins and RNA molecules that are kinetically trapped in misfolded states. Unassisted folding occurs by the kinetic partitioning mechanism according to which folding to the native state, with low probability as well as misfolding to one of the many metastable states, with high probability, occur rapidly. GroEL is an all‐purpose stochastic machine that assists misfolded substrate proteins to fold. The RNA chaperones such as CYT‐19, which are ATP‐consuming enzymes, help the folding of ribozymes that get trapped in metastable states for long times. GroEL does not interact with the folded proteins but CYT‐19 disrupts both the folded and misfolded ribozymes. The structures of GroEL and RNA chaperones are strikingly different. Despite these differences, the iterative annealing mechanism (IAM) quantitatively explains all the available experimental data for assisted folding of proteins and ribozymes. Driven by ATP binding and hydrolysis and GroES binding, GroEL undergoes a catalytic cycle during which it samples three allosteric states,T(apo),R(ATP bound), andR(ADP bound). Analyses of the experimental data show that the efficiency of the GroEL–GroES machinery and mutants is determined by the resetting ratekR ″  → T, which is largest for the wild‐type (WT) GroEL. Generalized IAM accurately predicts the folding kinetics ofTetrahymenaribozyme and its variants. Chaperones maximize the product of the folding rate and the steady‐state native state fold by driving the substrates out of equilibrium. Neither the absolute yield nor the folding rate is optimized.

    more » « less