The glucocorticoid receptor (GR) is a nuclear receptor that controls critical biological processes by regulating the transcription of specific genes. There is a known allosteric cross-talk between ...the ligand and coregulator binding sites within the GR ligand-binding domain that is crucial for the control of the functional response. However, the molecular mechanisms underlying such an allosteric control remain elusive. Here, molecular dynamics (MD) simulations, bioinformatic analysis, and biophysical measurements are integrated to capture the structural and dynamic features of the allosteric cross-talk within the GR. We identified a network of evolutionarily conserved residues that enables the allosteric signal transduction, in agreement with experimental data. MD simulations clarify how such a network is dynamically interconnected and offer a mechanistic explanation of how different peptides affect the intensity of the allosteric signal. This study provides useful insights to elucidate the GR allosteric regulation, ultimately providing a foundation for designing novel drugs.
Computer-aided drug design plays an important role in medicinal chemistry to obtain insights into molecular mechanisms and to prioritize design strategies. Although significant improvement has been ...made in structure based design, it still remains a key challenge to accurately model and predict induced fit mechanisms. Most of the current available techniques either do not provide sufficient protein conformational sampling or are too computationally demanding to fit an industrial setting. The current study presents a systematic and exhaustive investigation of predicting binding modes for a range of systems using PELE (Protein Energy Landscape Exploration), an efficient and fast protein–ligand sampling algorithm. The systems analyzed (cytochrome P, kinase, protease, and nuclear hormone receptor) exhibit different complexities of ligand induced fit mechanisms and protein dynamics. The results are compared with results from classical molecular dynamics simulations and (induced fit) docking. This study shows that ligand induced side chain rearrangements and smaller to medium backbone movements are captured well in PELE. Large secondary structure rearrangements, however, remain challenging for all employed techniques. Relevant binding modes (ligand heavy atom RMSD < 1.0 Å) can be obtained by the PELE method within a few hours of simulation, positioning PELE as a tool applicable for rapid drug design cycles.
Sensory rhodopsins are the primary receptors of vision in animals and phototaxis in microorganisms. Light triggers the rapid isomerization of a buried retinal chromophore, which the protein both ...accommodates and amplifies into the larger structural rearrangements required for signaling. We trapped an early intermediate of the photocycle of sensory rhodopsin II from
Natronobacterium pharaonis (pSRII) in 3D crystals and determined its X-ray structure to 2.3 Å resolution. The observed structural rearrangements were localized near the retinal chromophore, with a key water molecule becoming disordered and the retinal's β-ionone ring undergoing a prominent movement. Comparison with the early structural rearrangements of bacteriorhodopsin illustrates how modifications in the retinal binding pocket of pSRII allow subtle differences in the early relaxation of photoisomerized retinal.
In this study, we performed an extensive exploration of the ligand entry mechanism for members of the steroid nuclear hormone receptor family (androgen receptor, estrogen receptor α, glucocorticoid ...receptor, mineralocorticoid receptor, and progesterone receptor) and their endogenous ligands. The exploration revealed a shared entry path through the helix 3, 7, and 11 regions. Examination of the x-ray structures of the receptor-ligand complexes further showed two distinct folds of the helix 6–7 region, classified as “open” and “closed”, which could potentially affect ligand binding. To improve sampling of the helix 6–7 loop, we incorporated motion modes based on principal component analysis of existing crystal structures of the receptors and applied them to the protein-ligand sampling. A detailed comparison with the anisotropic network model (an elastic network model) highlights the importance of flexibility in the entrance region. While the binding (interaction) energy of individual simulations can be used to score different ligands, extensive sampling further allows us to predict absolute binding free energies and analyze reaction kinetics using Markov state models and Perron-cluster cluster analysis, respectively. The predicted relative binding free energies for three ligands binding to the progesterone receptor are in very good agreement with experimental results and the Perron-cluster cluster analysis highlighted the importance of a peripheral binding site. Our analysis revealed that the flexibility of the helix 3, 7, and 11 regions represents the most important factor for ligand binding. Furthermore, the hydrophobicity of the ligand influences the transition between the peripheral and the active binding site.
The glucocorticoid receptor (GR) is a ligand-activated transcription factor that binds DNA and assembles co-regulator complexes to regulate gene transcription. GR agonists are widely prescribed to ...people with inflammatory and autoimmune diseases. Here we present high-resolution, multidomain structures of GR in complex with ligand, DNA and co-regulator peptide. The structures reveal how the receptor forms an asymmetric dimer on the DNA and provide a detailed view of the domain interactions within and across the two monomers. Hydrogen-deuterium exchange and DNA-binding experiments demonstrate that ligand-dependent structural changes are communicated across the different domains in the full-length receptor. This study demonstrates how GR forms a distinct architecture on DNA and how signal transmission can be modulated by the ligand pharmacophore, provides a platform to build a new level of understanding of how receptor modifications can drive disease progression and offers key insight for future drug design.
Synthetic glucocorticoids (GC) are essential for the treatment of a broad range of inflammatory diseases. However, their use is limited by target related adverse effects on, e.g., glucose homeostasis ...and bone metabolism. Starting from a nonsteroidal GR ligand (4) that is a full agonist in reporter gene assays, we exploited key functional triggers within the receptor, generating a range of structurally diverse partial agonists. Of these, only a narrow subset exhibited full anti-inflammatory efficacy and a significantly reduced impact on adverse effect markers in human cell assays compared to prednisolone. This led to the discovery of AZD9567 (15) with excellent in vivo efficacy when dosed orally in a rat model of joint inflammation. Compound 15 is currently being evaluated in clinical trials comparing the efficacy and side effect markers with those of prednisolone.
The glucocorticoid receptor (GR) is a ligand-dependent transcription factor that plays a central role in inflammation. The GR activity is also modulated via protein–protein interactions, including ...binding of 14-3-3 proteins induced by GR phosphorylation. However, the specific phosphorylation sites on the GR that trigger these interactions and their functional consequences are less clear. Hence, we sought to examine this system in more detail. We used phosphorylated GR peptides, biophysical studies, and X-ray crystallography to identify key residues within the ligand-binding domain of the GR, T524 and S617, whose phosphorylation results in binding of the representative 14-3-3 protein 14-3-3ζ. A kinase screen identified misshapen-like kinase 1 (MINK1) as responsible for phosphorylating T524 and Rho-associated protein kinase 1 for phosphorylating S617; cell-based approaches confirmed the importance of both GR phosphosites and MINK1 but not Rho-associated protein kinase 1 alone in inducing GR–14-3-3 binding. Together our results provide molecular-level insight into 14-3-3-mediated regulation of the GR and highlight both MINK1 and the GR–14-3-3 axis as potential targets for future therapeutic intervention.
The ubiquitously expressed glucocorticoid receptor (GR) is a nuclear receptor that controls a broad range of biological processes and is activated by steroidal glucocorticoids such as hydrocortisone ...or dexamethasone. Glucocorticoids are used to treat a wide variety of conditions, from inflammation to cancer but suffer from a range of side effects that motivate the search for safer GR modulators. GR is also regulated outside the steroid-binding site through protein–protein interactions (PPIs) with 14-3-3 adapter proteins. Manipulation of these PPIs will provide insights into noncanonical GR signaling as well as a new level of control over GR activity. We report the first molecular glues that selectively stabilize the 14-3-3/GR PPI using the related nuclear receptor estrogen receptor α (ERα) as a selectivity target to drive design. These 14-3-3/GR PPI stabilizers can be used to dissect noncanonical GR signaling and enable the development of novel atypical GR modulators.
Aims
Corticosteroids are the treatment of choice for many inflammatory diseases but often lead to adverse effects, including hyperglycaemia. This study investigated the mechanisms driving ...differential effects on glucose control for AZD9567, an oral nonsteroidal selective glucocorticoid receptor modulator vs. prednisolone in 46 patients with type 2 diabetes mellitus.
Methods
In this randomized, double‐blind, 2‐way cross‐over study (NCT04556760), participants received either AZD9567 72 mg and prednisolone 40 mg daily (cohort 1); AZD9567 40 mg and prednisolone 20 mg daily (cohort 2); or placebo and prednisolone 5 mg daily (cohort 3). Treatment duration was 3 days with a 3‐week washout between treatment periods. Glycaemic control was assessed after a standardized meal and with continuous glucose monitoring.
Results
A significant difference between AZD9567 and prednisolone in favour of AZD9567 was observed for the change from baseline to Day 4 glucose excursions postmeal in cohort 1 (glucose area under the curve from 0 to 4 h −4.54%; 95% confidence interval CI: −8.88, −0.01; P = .049), but not in cohort 2 (−5.77%; 95% CI: −20.92, 12.29; P = .435). In cohort 1, significant differences between AZD9567 and prednisolone were also seen for the change from baseline to day 4 in insulin and glucagon secretion postmeal (P < .001 and P = .005, respectively) and change from baseline to Day 4 in GLP‐1 response (P = .022). Significant differences between AZD9567 and prednisolone for 24‐h glucose control were observed for both cohort 1 (−1.507 mmol/L; 95% CI: −2.0820, −0.9314; P < .001) and cohort 2 (−1.110 mmol/L; 95% CI −1.7257, −0.4941; P < .001).
Conclusion
AZD9567 significantly reduced treatment‐induced hyperglycaemia compared with prednisolone.
PDF
