scholarly journals Defining a canonical ligand-binding pocket in the orphan nuclear receptor Nurr1

2018 ◽  
Author(s):  
Ian Mitchelle S. de Vera ◽  
Paola Munoz-Tello ◽  
Venkatasubramanian Dharmarajan ◽  
David P. Marciano ◽  
Edna Matta-Camacho ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Nuclear Receptor ◽  
Unsaturated Fatty Acids ◽  
Solvent Accessibility ◽  
Physical Space ◽  
Binding Pocket ◽  
Orphan Nuclear Receptor ◽  
Ligand Binding Pocket ◽  
Exchange Mass Spectrometry ◽  
Dynamics Simulations

Nuclear receptor related 1 protein (Nurr1/NR4A2) is an orphan nuclear receptor that is considered to function without a canonical ligand-binding pocket. A crystal structure of the Nurr1 ligand-binding domain (LBD) revealed no physical space in the conserved region where other nuclear receptors with solvent accessible apo-protein ligand-binding pockets bind synthetic and natural ligands. Using solution NMR spectroscopy, hydrogen/deuterium exchange mass spectrometry, and molecular dynamics simulations, we show here that the putative canonical ligand-binding pocket in the Nurr1 LBD is dynamic with high solvent accessibility, exchanges between two or more conformations on the microsecond-to-millisecond timescale, and can expand from the collapsed crystalized conformation to allow binding of unsaturated fatty acids. These findings should stimulate future studies to probe the ligandability and druggability of Nurr1 for both endogenous and synthetic ligands, which could lead to new therapeutics for Nurr1-related diseases, including Parkinson’s disease and schizophrenia.

scholarly journals Defining a Canonical Ligand-Binding Pocket in the Orphan Nuclear Receptor Nurr1

Structure ◽  
2019 ◽  
Vol 27 (1) ◽  
pp. 66-77.e5 ◽  
Author(s):  
Ian Mitchelle S. de Vera ◽  
Paola Munoz-Tello ◽  
Jie Zheng ◽  
Venkatasubramanian Dharmarajan ◽  
David P. Marciano ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Nuclear Receptor ◽  
Binding Pocket ◽  
Orphan Nuclear Receptor ◽  
Ligand Binding Pocket

scholarly journals Defining a Ligand-Binding Pocket in the Orphan Nuclear Receptor Nurr1

2018 ◽  
Vol 114 (3) ◽  
pp. 66a
Author(s):  
Paola Munoz-Tello ◽  
Sarah Mosure ◽  
Patrick Griffin ◽  
Venkatasubramanian Dharmarajan ◽  
Ian de Vera ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Nuclear Receptor ◽  
Binding Pocket ◽  
Orphan Nuclear Receptor ◽  
Ligand Binding Pocket

scholarly journals SUN-LB138 Dynamic Structural Model of Testosterone Entry Into the Unliganded Androgen Receptor

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Irina Krylova ◽  
Fred J Schaufele ◽  
Christophe Guilbert
Keyword(s):  
Amino Acids ◽  
Androgen Receptor ◽  
Ligand Binding ◽  
Nuclear Receptors ◽  
Nuclear Receptor ◽  
Binding Pocket ◽  
Binding Domain ◽  
Receptor Ligand ◽  
Ligand Binding Pocket ◽  
Crystallographic Structures

Abstract Background: Crystallographic structures of nuclear receptor ligand binding domains provide a static model of a receptor stably wrapped around an internalized ligand. Understanding the dynamics of a receptor at different stages of ligand binding has been hampered by the paucity of crystal structures for unliganded nuclear receptors. Molecular dynamic models have been constructed for some nuclear receptors to fill that void. Methods: The molecular simulation docking program MORDOR (MOlecular Recognition with a Driven dynamics OptimizeR)(1) was used to study the structural dynamics of the androgen receptor ligand binding domain (AR LBD) modeled from the static structure of the AR LBD bound to testosterone (T) (PDB ID: 2AM9). The goals of the study were to understand a) the dynamic interaction of the T in its binding pocket, b) AR LBD structural flexibilities that permit T entry/exit from the binding pocket and c) a model of the unliganded AR LBD. Results: Modeling AR LBD structure flexibility over time revealed possible alternative dynamic structures, including those without ligand, overlaid against the canonical nuclear receptor structure. The model dynamically tracks the structural changes as a ligand enters into the ligand binding domain and nestles into the ligand binding pocket. The model predicted the appearance of alpha helices within the AR LBD that transiently fold/unfold during the ligand entry phases. Once in the pocket, the ligand itself remains very dynamic in a still flexible pocket. The model predicted also AR LBD amino acids that sequentially interact with the ligand during its dynamic entry into the AR LBD. Intriguingly, those AR amino acids include those mutated in castration-resistant prostate tumors that continue to grow during androgen suppression therapy. Functional studies showed those mutant ARs had a primary consequence of enhancing response to lower level T, and other androgens, consistent with their role in creating a higher affinity AR that can scavenge low-level androgens in an androgen-suppressed patient. Conclusions: The molecular model of T binding to the AR LBD suggests a degree of structural dynamism not evident in the crystallographic structures commonly associated with nuclear receptors. Some AR mutations activating prostate tumor growth may do so by impacting androgen entry/exit, rather than by altering androgen fit into the ligand binding pocket. Reference: (1) Guilbert C, James TL (2008) J Chem Inf Model. 2008 48(6): 1257-1268. doi: 10.1021/ci8000327

scholarly journals Kinetic and Thermodynamic Characterization of Dihydrotestosterone-Induced Conformational Perturbations in Androgen Receptor Ligand-Binding Domain

2009 ◽  
Vol 23 (8) ◽  
pp. 1231-1241 ◽  
Author(s):  
Ravi Jasuja ◽  
Jagadish Ulloor ◽  
Christopher M. Yengo ◽  
Karen Choong ◽  
Andrei Y. Istomin ◽  
...  
Keyword(s):  
Androgen Receptor ◽  
Ligand Binding ◽  
Conformational Changes ◽  
Solvent Accessibility ◽  
Molten Globule ◽  
Binding Pocket ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Second Derivative ◽  
Ligand Binding Pocket

Abstract Ligand-induced conformational perturbations in androgen receptor (AR) are important in coactivator recruitment and transactivation. However, molecular rearrangements in AR ligand-binding domain (AR-LBD) associated with agonist binding and their kinetic and thermodynamic parameters are poorly understood. We used steady-state second-derivative absorption and emission spectroscopy, pressure and temperature perturbations, and 4,4′-bis-anilinonaphthalene 8-sulfonate (bis-ANS) partitioning to determine the kinetics and thermodynamics of the conformational changes in AR-LBD after dihydrotestosterone (DHT) binding. In presence of DHT, the second-derivative absorption spectrum showed a red shift and a change in peak-to-peak distance. Emission intensity increased upon DHT binding, and center of spectral mass was blue shifted, denoting conformational changes resulting in more hydrophobic environment for tyrosines and tryptophans within a more compact DHT-bound receptor. In pressure perturbation calorimetry, DHT-induced energetic stabilization increased the Gibbs free energy of unfolding to 8.4 ± 1.3 kcal/mol from 3.5 ± 1.6 kcal/mol. Bis-ANS partitioning studies revealed that upon DHT binding, AR-LBD underwent biphasic rearrangement with a high activation energy (13.4 kcal/mol). An initial, molten globule-like burst phase (k ∼30 sec−1) with greater solvent accessibility was followed by rearrangement (k ∼0.01 sec−1), leading to a more compact conformation than apo-AR-LBD. Molecular simulations demonstrated unique sensitivity of tyrosine and tryptophan residues during pressure unfolding with rearrangement of residues in the coactivator recruitment surfaces distant from the ligand-binding pocket. In conclusion, DHT binding leads to energetic stabilization of AR-LBD domain and substantial rearrangement of residues distant from the ligand-binding pocket. DHT binding to AR-LBD involves biphasic receptor rearrangement including population of a molten globule-like intermediate state.

scholarly journals Identification of Kinase Inhibitors that Regulate Nuclear Receptor Nurr1 (NR4A2) Cellular Activity

2018 ◽  
Author(s):  
Kenya L. Williams ◽  
Carrow I. Wells ◽  
John T. Moore
Keyword(s):  
Ligand Binding ◽  
Nuclear Receptor ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Binding Pocket ◽  
Luciferase Reporter ◽  
Cellular Activity ◽  
Ligand Binding Pocket ◽  
Amino Terminal ◽  
Receptor Assays

AbstractThe ability to regulate the activity NR4A subfamily of nuclear receptors would be potentially useful in the treatment of multiple diseases, but to date, few regulators have been reported. This is likely due to the fact that the NR4A subfamily does not have a typical unoccupied nuclear receptor ligand binding pocket, but rather a pocket that is occupied with hydrophobic side chains of adjacent amino acids. It follows that traditional nuclear receptor assays that seek to identify ligands that bind within the ligand binding pocket would not be successful. We have thus focused on an alternate assay to identify NR4A regulators based on the fact that regulation of NR4A, at least partially, results from phosphorylation/dephosphorylation of the amino terminal region of the protein. We developed a medium throughput cellular assay using a fusion of the amino terminus of Nurr1 (NR4A2) with luciferase reporter and used the assay to screen a large and diverse protein kinase inhibitor set (PKIS). We identified multiple kinase inhibitor compounds from PKIS that significantly increased or decreased the cellular activity of Nurr1. These molecules serve as starting points to discover selective tools for regulation of Nurr1/Nurr1pathway.

scholarly journals Determinants of ligand selectivity in a cyclic nucleotide–regulated potassium channel

2014 ◽  
Vol 144 (1) ◽  
pp. 41-54 ◽  
Author(s):  
João Pessoa ◽  
Fátima Fonseca ◽  
Simone Furini ◽  
João H. Morais-Cabral
Keyword(s):  
Ligand Binding ◽  
Potassium Channel ◽  
Cyclic Nucleotide ◽  
Binding Pocket ◽  
Bound State ◽  
Triple Mutant ◽  
Ligand Binding Pocket ◽  
Ligand Selectivity ◽  
Molecular Determinants ◽  
Dynamics Simulations

Cyclic nucleotide–binding (CNB) domains regulate the activity of channels, kinases, exchange factors, and transcription factors. These proteins are highly variable in their ligand selectivity; some are highly selective for either cAMP or cGMP, whereas others are not. Several molecular determinants of ligand selectivity in CNB domains have been defined, but these do not provide a complete view of the selectivity mechanism. We performed a thorough analysis of the ligand-binding properties of mutants of the CNB domain from the MlotiK1 potassium channel. In particular, we defined which residues specifically favor cGMP or cAMP. Inversion of ligand selectivity, from favoring cAMP to favoring cGMP, was only achieved through a combination of three mutations in the ligand-binding pocket. We determined the x-ray structure of the triple mutant bound to cGMP and performed molecular dynamics simulations and a biochemical analysis of the effect of the mutations. We concluded that the increase in cGMP affinity and selectivity does not result simply from direct interactions between the nucleotide base and the amino acids introduced in the ligand-binding pocket residues. Rather, tighter cGMP binding over cAMP results from the polar chemical character of the mutations, from greater accessibility of water molecules to the ligand in the bound state, and from an increase in the structural flexibility of the mutated binding pocket.

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