In Science Journals.

Highlights from the family of journals.

Unlocking GPCR-ligand interactions: Measuring binding affinities with thermal shift assay.

G protein-coupled receptors (GPCRs) constitute the largest transmembrane protein superfamily, with over 800 representatives in the human genome. Recognized as pivotal targets in pharmacological research and drug discovery, these receptors play a crucial role in advancing therapeutics. Understanding the molecular mechanisms of receptor-ligand interactions is imperative for drug discovery applications.

Structural insights into the high basal activity and inverse agonism of the orphan receptor GPR6 implicated in Parkinson's disease.

GPR6 is an orphan G protein-coupled receptor with high constitutive activity found in D2-type dopamine receptor-expressing medium spiny neurons of the striatopallidal pathway, which is aberrantly hyperactivated in Parkinson's disease. Here, we solved crystal structures of GPR6 without the addition of a ligand (a pseudo-apo state) and in complex with two inverse agonists, including CVN424, which improved motor symptoms in patients with Parkinson's disease in clinical trials. In addition, we obtained a cryo-electron microscopy structure of the signaling complex between GPR6 and its cognate G heterotrimer.

Nicotinamide Riboside and CD38: Covalent Inhibition and Live-Cell Labeling.

Nicotinamide adenine dinucleotide (NAD) is required for a myriad of metabolic, signaling, and post-translational events in cells. Its levels in tissues and organs are closely associated with health conditions. The homeostasis of NAD is regulated by biosynthetic pathways and consuming enzymes.

Native mass spectrometry prescreening of G protein-coupled receptor complexes for cryo-EM structure determination.

G protein-coupled receptors (GPCRs) are essential transmembrane proteins playing key roles in human health and disease. Understanding their atomic-level molecular structure and conformational states is imperative for advancing drug development. Recent breakthroughs in single-particle cryogenic electron microscopy (cryo-EM) have propelled the structural biology of GPCRs into a new era.

Insulator-based dielectrophoresis-assisted separation of insulin secretory vesicles.

Organelle heterogeneity and inter-organelle contacts within a single cell contribute to the limited sensitivity of current organelle separation techniques, thus hindering organelle subpopulation characterization. Here, we use direct current insulator-based dielectrophoresis (DC-iDEP) as an unbiased separation method and demonstrate its capability by identifying distinct distribution patterns of insulin vesicles from INS-1E insulinoma cells. A multiple voltage DC-iDEP strategy with increased range and sensitivity has been applied, and a differentiation factor (ratio of electrokinetic to dielectrophoretic mobility) has been used to characterize features of insulin vesicle distribution patterns.

Functional GPCR Expression in Eukaryotic LEXSY System.

G protein-coupled receptors (GPCRs) form the largest superfamily of membrane proteins in the human genome, and represent one of the most important classes of drug targets. Their structural studies facilitate rational drug discovery. However, atomic structures of only about 20% of human GPCRs have been solved to date.

Structural diversity of leukotriene G-protein coupled receptors.

Dihydroxy acid leukotriene (LTB) and cysteinyl leukotrienes (LTC, LTD, and LTE) are inflammatory mediators derived from arachidonic acid via the 5-lipoxygenase pathway. While structurally similar, these two types of leukotrienes (LTs) exert their functions through interactions with two distinct G protein-coupled receptor (GPCR) families, BLT and CysLT receptors, which share low sequence similarity and belong to phylogenetically divergent GPCR groups. Selective antagonism of LT receptors has been proposed as a promising strategy for the treatment of many inflammation-related diseases including asthma and chronic obstructive pulmonary disease, rheumatoid arthritis, cystic fibrosis, diabetes, and several types of cancer.

MicroED structure of the human vasopressin 1B receptor.

The small size and flexibility of G protein-coupled receptors (GPCRs) have long posed a significant challenge to determining their structures for research and therapeutic applications. Single particle cryogenic electron microscopy (cryoEM) is often out of reach due to the small size of the receptor without a signaling partner. Crystallization of GPCRs in lipidic cubic phase (LCP) often results in crystals that may be too small and difficult to analyze using X-ray microcrystallography at synchrotron sources or even serial femtosecond crystallography at X-ray free electron lasers.

Structural insights into angiotensin receptor signaling modulation by balanced and biased agonists.

The peptide hormone angiotensin II regulates blood pressure mainly through the type 1 angiotensin II receptor AT R and its downstream signaling proteins G and β-arrestin. AT R blockers, clinically used as antihypertensive drugs, inhibit both signaling pathways, whereas AT R β-arrestin-biased agonists have shown great potential for the treatment of acute heart failure. Here, we present a cryo-electron microscopy (cryo-EM) structure of the human AT R in complex with a balanced agonist, Sar -AngII, and G protein at 2.

The activation mechanism and antibody binding mode for orphan GPR20.

GPR20 is a class-A orphan G protein-coupled receptor (GPCR) and a potential therapeutic target for gastrointestinal stromal tumors (GIST) owing to its differentially high expression. An antibody-drug conjugate (ADC) containing a GPR20-binding antibody (Ab046) was recently developed in clinical trials for GIST treatment. GPR20 constitutively activates Gi proteins in the absence of any known ligand, but it remains obscure how this high basal activity is achieved.

Serial femtosecond crystallography.

With the advent of X-ray Free Electron Lasers (XFELs), new, high-throughput serial crystallography techniques for macromolecular structure determination have emerged. Serial femtosecond crystallography (SFX) and related methods provide possibilities beyond canonical, single-crystal rotation crystallography by mitigating radiation damage and allowing time-resolved studies with unprecedented temporal resolution. This primer aims to assist structural biology groups with little or no experience in serial crystallography planning and carrying out a successful SFX experiment.

GPCR Agonist-to-Antagonist Conversion: Enabling the Design of Nucleoside Functional Switches for the A Adenosine Receptor.

Modulators of the G protein-coupled A adenosine receptor (AAR) have been considered promising agents to treat Parkinson's disease, inflammation, cancer, and central nervous system disorders. Herein, we demonstrate that a thiophene modification at the C8 position in the common adenine scaffold converted an AAR agonist into an antagonist. We synthesized and characterized a novel AAR antagonist, (LJ-4517), with = 18.

Structural basis for receptor selectivity and inverse agonism in S1P receptors.

The bioactive lysophospholipid sphingosine-1-phosphate (S1P) acts via five different subtypes of S1P receptors (S1PRs) - S1P. S1P is predominantly expressed in nervous and immune systems, regulating the egress of natural killer cells from lymph nodes and playing a role in immune and neurodegenerative disorders, as well as carcinogenesis. Several S1PR therapeutic drugs have been developed to treat these diseases; however, they lack receptor subtype selectivity, which leads to side effects.

High ligand efficiency quinazoline compounds as novel A adenosine receptor antagonists.

The past fifty years have been marked by the surge of neurodegenerative diseases. Unfortunately, current treatments are only symptomatic. Hence, the search for new and innovative therapeutic targets for curative treatments becomes a major challenge.

Inhibition of the angiotensin II type 2 receptor ATR is a novel therapeutic strategy for glioblastoma.

Glioblastoma (GBM) is an aggressive malignant primary brain tumor with limited therapeutic options. We show that the angiotensin II (AngII) type 2 receptor (ATR) is a therapeutic target for GBM and that AngII, endogenously produced in GBM cells, promotes proliferation through ATR. We repurposed EMA401, an ATR antagonist originally developed as a peripherally restricted analgesic, for GBM and showed that it inhibits the proliferation of ATR-expressing GBM spheroids and blocks their invasiveness and angiogenic capacity.

Crystallization of Microbial Rhodopsins.

Microbial rhodopsins are light-sensitive transmembrane proteins, evolutionary adapted by various organisms like archaea, bacteria, simple eukaryote, and viruses to utilize solar energy for their survival. A complete understanding of functional mechanisms of these proteins is not possible without the knowledge of their high-resolution structures, which can be primarily obtained by X-ray crystallography. This technique, however, requires high-quality crystals, growing of which is a great challenge especially in case of membrane proteins.

In Science Journals.

Highlights from the family of journals.

Structural basis of GABA reuptake inhibition.

γ-Aminobutyric acid (GABA) transporter 1 (GAT1) regulates neuronal excitation of the central nervous system by clearing the synaptic cleft of the inhibitory neurotransmitter GABA upon its release from synaptic vesicles. Elevating the levels of GABA in the synaptic cleft, by inhibiting GABA reuptake transporters, is an established strategy to treat neurological disorders, such as epilepsy. Here we determined the cryo-electron microscopy structure of full-length, wild-type human GAT1 in complex with its clinically used inhibitor tiagabine, with an ordered part of only 60 kDa.