Jana Selent

Induced Effects of Sodium Ions on Dopaminergic G-Protein Coupled Receptors

G-protein coupled receptors, the largest family of proteins in the human genome, are involved in many complex signal transduction pathways, typically activated by orthosteric ligand binding and subject to allosteric modulation. Dopaminergic receptors, belonging to the class A family of G-protein coupled receptors, are known to be modulated by sodium ions from an allosteric binding site, although the details of sodium effects on the receptor have not yet been described. In an effort to understand these effects, we performed microsecond scale all-atom molecular dynamics simulations on the dopaminergic D2 receptor, finding that sodium ions enter the receptor from the extracellular side and bind at a deep allosteric site (Asp2.50). Remarkably, the presence of a sodium ion at this allosteric site induces a conformational change of the rotamer toggle switch Trp6.48 which locks in a conformation identical to the one found in the partially inactive state of the crystallized human β2 adrenergic receptor. This study provides detailed quantitative information about binding of sodium ions in the D2 receptor and reports a possibly important sodium-induced conformational change for modulation of D2 receptor function.

Multi-Receptor Binding Profile of Clozapine and Olanzapine: A Structural Study Based on the New β2 Adrenergic Receptor Template

Schizophrenia is a devastating mental disorder that has a large impact on the quality of life for those who are afflicted and is very costly for families and society.[1] Although the etiology of schizophrenia is still unknown and no cure has yet been found, it is treatable, and pharmacological therapy often produces satisfactory results. Among the various antipsychotic drugs in use, clozapine is widely recognized as one ofthe most clinically effective agents, even if it elicits significant side effects such as metabolic disorders and agranulocytosis. Clozapine and the closely related compound olanzapine are good examples ofdrug s with a complex multi-receptor profile ;[2] they have affinities toward serotonin, dopamine, a adrenergic, muscarinic, and histamine receptors, among others.

Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease.

Equine Estrogens Impair Nitric Oxide Production and Endothelial Nitric Oxide Synthase Transcription in Human Endothelial Cells Compared With the Natural 17β-Estradiol

Conjugated equine estrogen therapy is the most common hormone replacement strategy used to treat postmenopausal women. However, the ability of an individual conjugated equine estrogen to modulate NO production and, therefore, to induce cardiovascular protection is largely unknown. The effects of equine and naturally occurring estrogens on NO generation were evaluated in human aortic endothelial cells by measuring in vivo NO production, as well as NO synthase (eNOS) activity and expression. The transcriptional activity on the eNOS gene was determined by the ability of estrogen receptors (&agr; and &bgr;) to activate the eNOS promoter and induce transcription. Docking and molecular dynamics simulations were used to study structural features of the interaction between estrogenic compounds and estrogen receptor-&agr;. After 24 hours of incubation, we found that estrone upregulated NO production almost as effectively as estradiol by increasing eNOS activity and expression. However, the effect of equine estrogens (equilin, equilenin, and their metabolites) were marked decreased. eNOS promoter activity by equine estrogens was 30% to 50% lower than the naturally occurring estrogens. Computational analysis of estrogen molecules revealed that position 17 and the saturation of estrogenic compounds in ring B are important determinants for estrogen receptor-&agr; transcriptional activity. Equine estrogens increase NO production less effectively than naturally occurring estrogens, partially because of their lesser ability to activate the eNOS promoter and induce transcription. Differences in NO production by different estrogens may account for the differences in cardiovascular benefits achieved by the distinct estrogen replacement therapies.

Membrane Protein Structure, Function and Dynamics: A Perspective from Experiments and Theory

Membrane proteins mediate processes that are fundamental for the flourishing of biological cells. Membrane-embedded transporters move ions and larger solutes across membranes; receptors mediate communication between the cell and its environment and membrane-embedded enzymes catalyze chemical reactions. Understanding these mechanisms of action requires knowledge of how the proteins couple to their fluid, hydrated lipid membrane environment. We present here current studies in computational and experimental membrane protein biophysics, and show how they address outstanding challenges in understanding the complex environmental effects on the structure, function, and dynamics of membrane proteins.

Synthesis, binding affinity and SAR of new benzolactam derivatives as dopamine D3 receptor ligands

A series of new benzolactam derivatives was synthesized and the derivatives were evaluated for their affinities at the dopamine D(1), D(2), and D(3) receptors. Some of these compounds showed high D(2) and/or D(3) affinity and selectivity over the D(1) receptor. The SAR study of these compounds revealed structural characteristics that decisively influenced their D(2) and D(3) affinities. Structural models of the complexes between some of the most representative compounds of this series and the D(2) and D(3) receptors were obtained with the aim of rationalizing the observed experimental results. Moreover, selected compounds showed moderate binding affinity on 5-HT(2A) which could contribute to reducing the occurrence of extrapyramidal side effects as potential antipsychotics.

Progress in the structural prediction of G protein-coupled receptors: D3 receptor in complex with eticlopride.

Predicting the three‐dimensional structure of ligand–receptor complexes involving G protein‐coupled receptors (GPCRs) is still a challenging task in rational drug design. To evaluate the reliability of the GPCR structural prediction, only a couple of community‐wide assessments have been carried out. Our participation in the last edition, DOCK2010, involved the blind prediction of the dopaminergic D3 receptor in complex with the D2/D3 selective antagonist eticlopride for which the crystal structure has been recently released. Here, we describe a methodology that succeeded to produce a correctly predicted eticlopride‐D3 receptor complex out of three submitted models. Ranking the obtained models in the correct order is the main challenge due to subtle structural differences in the complex that are not sufficiently captured by conventional scoring functions. Importantly, our work reveals that a correct ranking is obtained by including a more sophisticated description of conformational ligand energy on binding. All in all, this case study highlights the current progress in modeling GPCR complexes and underlines that in silico modeling can be a valuable complement in GPCR drug discovery. Proteins 2011;

Novel Insights into Biased Agonism at G Protein-Coupled Receptors and their Potential for Drug Design

G-protein coupled receptors (GPCRs) are the most important class of current pharmacological targets. However, it is now widely acknowledged that their regulation is more complex than previously thought: the evidence that GPCRs can couple to several effector pathways, and the existence of biased agonists able to activate them differentially, has introduced a new level of complexity in GPCR drug research. Considering bias represents a challenge for the research of new GPCR modulators, because it demands a detailed characterization of compound properties for several effector pathways. Still, biased ligands could provide an opportunity to modulate GPCR function in a finer way and to separate therapeutic from side effects. Nowadays, a variety of agonists for GPCRs have been described, which differ in their ability to promote receptor coupling to different Gprotein families or even subunits, recruit signal transducers such as arrestins, activate a variety of downstream molecular pathways and induce certain phosphorylation signatures or gene expression patterns. In this review, we will cover some of the experimental techniques currently used to understand and characterize biased agonism and discuss their strengths and limitations. Additionally, we will comment on the computational efforts that are being devoted to study ligand-induced bias and on the potential they hold for rationalizing its structural determinants. Finally, we will discuss which of these strategies could be used for the rational design of biased ligands and give some examples of the potential therapeutic value of this class of compounds.

Synthesis, Binding Affinity, and Molecular Docking Analysis of New Benzofuranone Derivatives as Potential Antipsychotics

The complex etiology of schizophrenia has prompted researchers to develop clozapine-related multitarget strategies to combat its symptoms. Here we describe a series of new 6-aminomethylbenzofuranones in an effort to find new chemical structures with balanced affinities for 5-HT2 and dopamine receptors. Through biological and computational studies of 5-HT2A and D2 receptors, we identified the receptor serine residues S3.36 and S5.46 as the molecular keys to explaining the differences in affinity and selectivity between these new compounds for this group of receptors. Specifically, the ability of these compounds to establish one or two H-bonds with these key residues appears to explain their difference in affinity. In addition, we describe compound 2 (QF1004B) as a tool to elucidate the role of 5-HT2C receptors in mediating antipsychotic effects and metabolic adverse events. The compound 16a (QF1018B) showed moderate to high affinities for D2 and 5-HT2A receptors, and a 5-HT2A/D2 ratio was predictive of an atypical antipsychotic profile.

Membrane-Sensitive Conformational States of Helix 8 in the Metabotropic Glu2 Receptor, a Class C GPCR

The recent elucidation of the X-ray structure of several class A GPCRs clearly indicates that the amphipathic helix 8 (H8) is a conserved structural domain in most crystallized GPCRs. Very little is known about the presence and the possible role of an analogous H8 domain in the distantly related class C GPCRs. In this study, we investigated the structural properties for the H8 domain of the mGluR2 receptor, a class C GPCR, by applying extended molecular dynamics simulations. Our study indicates that the amphipathic H8 adopts membrane-sensitive conformational states, which depend on the membrane composition. Cholesterol-rich membranes stabilize the helical structure of H8 whereas cholesterol-depleted membranes induce a disruption of H8. The observed link between membrane cholesterol levels and H8 conformational states suggests that H8 behaves as a sensor of cholesterol concentration.