50 Years of Image Analysis

In the 10 years since our previous International Union of Basic and Clinical Pharmacology report on the nomenclature and classification of adenosine receptors, no developments have led to major changes in the recommendations. However, there have been so many other developments that an update is needed. The fact that the structure of one of the adenosine receptors has recently been solved has already led to new ways of in silico screening of ligands. The evidence that adenosine receptors can form homo- and heteromultimers has accumulated, but the functional significance of such complexes remains unclear. The availability of mice with genetic modification of all the adenosine receptors has led to a clarification of the functional roles of adenosine, and to excellent means to study the specificity of drugs. There are also interesting associations between disease and structural variants in one or more of the adenosine receptors. Several new selective agonists and antagonists have become available. They provide improved possibilities for receptor classification. There are also developments hinting at the usefulness of allosteric modulators. Many drugs targeting adenosine receptors are in clinical trials, but the established therapeutic use is still very limited.

https://pharmrev.aspetjournals.org/content/pharmrev/63/1/1.full.pdf

International Union of Basic and Clinical Pharmacology. LXXXI. Nomenclature and Classification of Adenosine Receptors—An Update

During the past few years, crystallography of G protein–coupled receptors (GPCRs) has experienced exponential growth, resulting in the determination of the structures of 16 distinct receptors—9 of them in 2012 alone. Including closely related subtype homology models, this coverage amounts to approximately 12% of the human GPCR superfamily. The adrenergic, rhodopsin, and adenosine receptor systems are also described by agonist-bound active-state structures, including a structure of the receptor–G protein complex for the β2-adrenergic receptor. Biochemical and biophysical techniques, such as nuclear magnetic resonance and hydrogen-deuterium exchange coupled with mass spectrometry, are providing complementary insights into ligand-dependent dynamic equilibrium between different functional states. Additional details revealed by high-resolution structures illustrate the receptors as allosteric machines that are controlled not only by ligands but also by ions, lipids, cholesterol, and water. This wealth of data ...

Structure-Function of the G Protein–Coupled Receptor Superfamily

G protein-coupled receptors (GPCRs) represent the largest family of cell-surface receptors. These receptors are natural allosteric proteins because agonist-mediated signaling by GPCRs requires a conformational change in the receptor protein transmitted between two topographically distinct binding sites, one for the agonist and another for the G protein. It is now becoming increasingly recognized, however, that the agonist-bound GPCR can also form ternary complexes with other ligands or “accessory” proteins and display altered binding and/or signaling properties in relation to the binary agonist-receptor complex. Allosteric sites on GPCRs represent novel drug targets because allosteric modulators possess a number of theoretical advantages over classic orthosteric ligands, such as a ceiling level to the allosteric effect and a potential for greater GPCR subtype-selectivity. Because of the noncompetitive nature of allosteric phenomena, the detection and quantification of such effects often relies on a combination of equilibrium binding, nonequilibrium kinetic, and functional signaling assays. This review discusses the development and properties of allosteric receptor models for GPCRs and the detection and quantification of allosteric effects. Moreover, we provide an overview of the current knowledge regarding the location of possible allosteric sites on GPCRs and candidate endogenous allosteric modulators. Finally, we discuss the potential for allosteric effects arising from the formation of GPCR oligomers or GPCRs complexed with accessory cellular proteins. It is proposed that the study of allosteric phenomena will become of progressively greater import to the drug discovery process due to the advent of newer and more sensitive GPCR screening technologies.

/pdf/g-protein-coupled-receptor-allosterism-and-complexing-3fkjazohf8.pdf

G Protein-Coupled Receptor Allosterism and Complexing

Organisms of all domains of life use photoreceptor proteins to sense and respond to light. The light-sensitivity of photoreceptor proteins arises from bound chromophores such as retinal in retinylidene proteins, bilin in biliproteins, and flavin in flavoproteins. Rhodopsins found in Eukaryotes, Bacteria, and Archaea consist of opsin apoproteins and a covalently linked retinal which is employed to absorb photons for energy conversion or the initiation of intra- or intercellular signaling.1 Both functions are important for organisms to survive and to adapt to the environment. While lower organisms utilize the family of microbial rhodopsins for both purposes, animals solely use a different family of rhodopsins, a specialized subset of G-protein-coupled receptors (GPCRs).1,2 Animal rhodopsins, for example, are employed in visual and nonvisual phototransduction, in the maintenance of the circadian clock and as photoisomerases.3,4 While sharing practically no sequence similarity, microbial and animal rhodopsins, also termed type-I and type-II rhodopsins, respectively, share a common architecture of seven transmembrane α-helices (TM) with the N- and C-terminus facing out- and inside of the cell, respectively (Figure ​(Figure11).1,5 Retinal is attached by a Schiff base linkage to the e-amino group of a lysine side chain in the middle of TM7 (Figures ​(Figures11 and ​and2).2). The retinal Schiff base (RSB) is protonated (RSBH+) in most cases, and changes in protonation state are integral to the signaling or transport activity of rhodopsins.



Figure 1

Topology of the retinal proteins. (A) These membrane proteins contain seven α-helices (typically denoted helix A to G in microbial opsins and TM1 to 7 in the animal opsins) spanning the lipid bilayer. The N-terminus faces the outside of the cell ...

Microbial and animal rhodopsins: structures, functions, and molecular mechanisms.

Efficient Coupling of Transducin to Monomeric Rhodopsin in a Phospholipid Bilayer

G protein-coupled receptor (GPCR)-mediated signal transduction has been studied for more than a century. Despite the intense focus on this class of proteins, a molecular understanding of what constitutes the functional form of the receptor is still uncertain. GPCRs have traditionally been conceptualized as monomeric proteins, and this view has changed little over the years until relatively recently. Recent biochemical and biophysical studies have challenged this traditional concept, and point instead to a mechanistic view of signal transduction wherein the receptor functions as an oligomer. Cooperative interactions within such an oligomeric array may be critical for the propagation of an external signal across the cell membrane and to the G protein, and may therefore underlie the mechanistic basis of signaling.

/pdf/oligomerization-of-g-protein-coupled-receptors-past-present-297y78ohgh.pdf

Oligomerization of G protein-coupled receptors: past, present, and future.

The rod outer segment (ROS) of photoreceptor cells houses all components necessary for phototransduction, a set of biochemical reactions that amplify and propagate a light signal. Theoretical approaches to quantify this process require precise information about the physical boundaries of the ROS. Dimensions of internal structures within the ROS of mammalian species have yet to be determined with the precision required for quantitative considerations. Cryoelectron tomography was utilized to obtain reliable three-dimensional morphological information about this important structure from murine retina. Vitrification of samples permitted imaging of the ROS in a minimally perturbed manner and the preservation of substructures. Tomograms revealed the characteristic highly organized arrangement of disc membranes stacked on top of one another with a surrounding plasma membrane. Distances among the various membrane components of the ROS were measured to define the space available for phototransduction to occur. Reconstruction of segments of the ROS from single-axis tilt series images provided a glimpse into the three-dimensional architecture of this highly differentiated neuron. The reconstructions revealed spacers that likely maintain the proper distance between adjacent discs and between discs and the plasma membrane. Spacers were found distributed throughout the discs, including regions that are distant from the rim region of discs.

https://rupress.org/jcb/article-pdf/177/5/917/1330267/jcb_200612010.pdf

Three-dimensional architecture of murine rod outer segments determined by cryoelectron tomography

Transformation of G protein–coupled receptors (GPCRs) from a quiescent to an active state initiates signal transduction. All GPCRs share a common architecture comprising seven transmembranespanning α-helices, which accommodates signal propagation from a diverse repertoire of external stimuli across biological membranes to a heterotrimeric G protein. Signal propagation through the transmembrane helices likely involves mechanistic features common to all GPCRs. The structure of the light receptor rhodopsin may serve as a prototype for the transmembrane architecture of GPCRs. Early biochemical, biophysical, and pharmacological studies led to the conceptualization of receptor activation based on the context of two-state equilibrium models and conformational changes in protein structure. More recent studies indicate a need to move beyond these classical paradigms and to consider additional aspects of the molecular character of GPCRs, such as the oligomerization and dynamics of the receptor.

Paul S.-H. Park

Papers

Efficient Coupling of Transducin to Monomeric Rhodopsin in a Phospholipid Bilayer

Oligomerization of G protein-coupled receptors: past, present, and future.

Three-dimensional architecture of murine rod outer segments determined by cryoelectron tomography

Activation of G Protein-Coupled Receptors: Beyond Two-State Models and Tertiary Conformational Changes

Atomic force microscopy: a multifaceted tool to study membrane proteins and their interactions with ligands.