Plasmon Resonance Studies of Agonist/Antagonist Binding to theHuman δ-Opioid Receptor: New Structural Insights into Receptor-Ligand Interactions

Salamon, Zdzislaw; Cowell, Scott; Varga, Éva; Yamamura, Henry I.; Hruby, Victor J.; Tollin, Gordon

doi:10.1016/s0006-3495(00)76489-7

Cited by 103 publications

(126 citation statements)

References 30 publications

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“…We have observed the same slow rate of agonist-induced conformational change with a different f luorescent reporter on Cys-125 in TM3 and on Cys-285 in TM6 of the ␤ 2 AR (Fig. 1 A) (23), and Salamon et al observed a similar rate of agonistinduced conformational changes in the ␦-opioid receptor analyzed by surface plasmon resonance spectroscopy (24). Thus, agonist binding precedes the conformational change.…”

Section: The Effect Of Full and Partial Agonists On The Fluorescence supporting

confidence: 69%

Agonist-induced conformational changes in the G-protein-coupling domain of the β ₂ adrenergic receptor

Ghanouni

Steenhuis

Farrens

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

314

271

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The majority of extracellular physiologic signaling molecules act by stimulating GTP-binding protein (G-protein)-coupled receptors (GPCRs). To monitor directly the formation of the active state of a prototypical GPCR, we devised a method to site specifically attach fluorescein to an endogenous cysteine (Cys-265) at the cytoplasmic end of transmembrane 6 (TM6) of the ␤2 adrenergic receptor (␤2AR), adjacent to the G-protein-coupling domain. We demonstrate that this tag reports agonist-induced conformational changes in the receptor, with agonists causing a decline in the fluorescence intensity of fluorescein-␤ 2AR that is proportional to the biological efficacy of the agonist. We also find that agonists alter the interaction between the fluorescein at Cys-265 and fluorescence-quenching reagents localized to different molecular environments of the receptor. These observations are consistent with a rotation and͞or tilting of TM6 on agonist activation. Our studies, when compared with studies of activation in rhodopsin, indicate a general mechanism for GPCR activation; however, a notable difference is the relatively slow kinetics of the conformational changes in the ␤2AR, which may reflect the different energetics of activation by diffusible ligands.D espite diverse physiologic roles, the majority of GTPbinding protein (G-protein)-coupled receptors (GPCRs) are thought to share a common activation mechanism. Briefly, agonists induce conformational changes in receptors, which then stimulate heterotrimeric G proteins. Activated G proteins influence cellular physiology by modulating specific effector enzymes and ion channels involved in cardiovascular, neural, endocrine, and sensory signaling systems (1). GPCRs share a common structural motif consisting of seven TM helices with an extracellular amino terminus. For all known GPCRs, the site of ligand binding is distant from the site of G-protein regulation (2). Therefore, the overall structural effects of agonists binding to extracellular sequences or TM domains must physically converge at the cytoplasmic interface between the receptor and its G protein.We chose to characterize the conformational changes involved in G-protein activation by studying the human ␤ 2 adrenergic receptor (␤ 2 AR). The ␤ 2 AR is an important pharmaceutical target for pulmonary and cardiovascular diseases, and a wide spectrum of pharmacologically well-characterized agonists and antagonists is readily available (3). Moreover, extensive mutagenesis studies have identified amino acids involved in ligand binding and G-protein coupling (Fig. 1 A), thereby providing a basis for focusing our studies to domains likely to report conformational changes involved in receptor activation (4). We devised a means of labeling purified detergent-solubilized ␤ 2 AR at Cys-265 in the carboxyl-terminal region of IC3 with the sulfhydryl-reactive f luorescent probe f luorescein maleimide (FM). Cys-265 is found in the native receptor, and labeling at this position did not require alteration of the receptor's amino acid se...

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Section: The Effect Of Full and Partial Agonists On The Fluorescence supporting

confidence: 69%

Agonist-induced conformational changes in the G-protein-coupling domain of the β ₂ adrenergic receptor

Ghanouni

Steenhuis

Farrens

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

314

271

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“…Unexpectedly, the interaction resulted in a negative curve. The generation of negative curves has previously been linked to a ligand-induced structural change; for example, the immobilized component may undergo a decrease of its hydrodynamic radius (29,30). It is plausible that the negative curve obtained here is due to a heparin-induced conformational change of the lipoprotein.…”

Section: Hs/heparinmentioning

confidence: 60%

“…Negative signals can be observed when the immobilized ligand undergoes a conformational change, often related to an analyte-induced decrease of its hydrodynamic radius. This change in conformation results in a decrease of the refractive index (29,30). Thus, the negative signals observed for HS/heparin interaction with HDL-SAA likely correspond to a conformational change of the lipoprotein.…”

Section: Discussionmentioning

confidence: 95%

Heparan Sulfate Dissociates Serum Amyloid A (SAA) from Acute-phase High-density Lipoprotein, Promoting SAA Aggregation

Noborn

Ancsin

Ubhayasekera

et al. 2012

Journal of Biological Chemistry

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Background: Serum amyloid A (SAA) is normally associated with the high-density lipoprotein (HDL). Results: Heparan sulfate (HS) dissociates SAA from HDL, leading to AA amyloidosis. The activity requires a minimum length of 12-14 sugar units. Conclusion:The result proposes an explanation for the findings that short HS precludes AA amyloidosis. Significance: This study defines a novel role for HS in AA amyloidosis.

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“…Conversely, they repulse if they present a different mismatch sign [101]. For instance, the activation of a receptor after ligand binding can change its hydrophobic thickness [102] and consequently the interaction potential with neighbouring proteins.…”

Section: Appendix 2 Nature Of Protein-protein Interactionsmentioning

confidence: 99%

What do diffusion measurements tell us about membrane compartmentalisation? Emergence of the role of interprotein interactions

2008

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The techniques of diffusion analysis based on optical microscopy approaches have revealed a great diversity of the dynamic organisation of cell membranes. For a long period, two frameworks have dominated the way of representing the membrane structure: the membrane skeleton fences and the lipid raft models. Progresses in the methods of data analysis have shed light on the features and consequently the possible origin of membrane domains: Inter-protein interactions play a role in confinement. Innovative developments pushing forward the spatiotemporal resolution limits are currently emerging, which are likely to provide in the future a detailed understanding of the intimate functional dynamic organisation of the cell membrane. Keywords Membrane domain . Diffusion . Protein interaction OverviewThe main function of membranes is to generate close compartments: The cell itself (by the plasma membrane) and also the nucleus (by nuclear envelope), the Golgi apparatus, the endoplasmic reticulum, various organelles or the vesicles which are involved in transport processes. Membranes delimit these structures and allow them to have a different composition from the rest of the cell by restricting the diffusion of ions and macromolecules. The specific transport of molecules or information across the membrane is devoted to membrane proteins. Membranes are essentially composed of lipid and proteins, each of them representing about 50% in weight. Lipids are amphiphilic molecules that spontaneously form a bilayer in water. Among the variety of lipids, cholesterol has a specific place since it is particularly abundant in eukaryotic cells. Cholesterol can modify the lipid molecular order [1] and is presumed to participate in lipid micro-phase separation (rafts) [2][3][4][5].Since the structure of biological membranes is maintained by non-covalent bonds, they have first been considered as a fluid lipid bilayer in which embedded proteins are free to diffuse [6]. After the advent of this fluid mosaic model postulating a random distribution of membrane molecules [6], experimental evidence showed that the proteins and lipids are distributed heterogeneously in the membrane. For example, incomplete recoveries were systematically observed in fluorescence recovery after photobleaching (FRAP; see "Appendix 1") experiments. The first indication of the presence of micrometer-sized domains was obtained by FRAP. Yechiel and Edidin found a decrease of the mobile fraction with increasing radius of the bleaching spot ([7, 8] and see "Appendix 1").The huge diversity of lipids and proteins implies that a very large number of combinatorial interactions can occur between them [9]. Since all lipids and proteins do not J Chem Biol (2008) [14,15]. Interactions of membrane proteins with the cytoskeleton or with the glycocalyx can also affect the membrane organisation and are likely to play a confining role [16]. A significant challenge of cell biology is to characterise and to understand not only the origin but also the role of these micrometric ...

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Plasmon Resonance Studies of Agonist/Antagonist Binding to theHuman δ-Opioid Receptor: New Structural Insights into Receptor-Ligand Interactions

Cited by 103 publications

References 30 publications

Agonist-induced conformational changes in the G-protein-coupling domain of the β ₂ adrenergic receptor

Agonist-induced conformational changes in the G-protein-coupling domain of the β ₂ adrenergic receptor

Heparan Sulfate Dissociates Serum Amyloid A (SAA) from Acute-phase High-density Lipoprotein, Promoting SAA Aggregation

What do diffusion measurements tell us about membrane compartmentalisation? Emergence of the role of interprotein interactions

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Plasmon Resonance Studies of Agonist/Antagonist Binding to theHuman δ-Opioid Receptor: New Structural Insights into Receptor-Ligand Interactions

Cited by 103 publications

References 30 publications

Agonist-induced conformational changes in the G-protein-coupling domain of the β 2 adrenergic receptor

Agonist-induced conformational changes in the G-protein-coupling domain of the β 2 adrenergic receptor

Heparan Sulfate Dissociates Serum Amyloid A (SAA) from Acute-phase High-density Lipoprotein, Promoting SAA Aggregation

What do diffusion measurements tell us about membrane compartmentalisation? Emergence of the role of interprotein interactions

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Agonist-induced conformational changes in the G-protein-coupling domain of the β ₂ adrenergic receptor

Agonist-induced conformational changes in the G-protein-coupling domain of the β ₂ adrenergic receptor