Crystal structure of a common GPCR-binding interface for G protein and arrestin

Szczepek, Michal; Beyrière, Florent; Hofmann, Klaus; Elgeti, Matthias; Kazmin, Roman; Rose, Alexander; Bartl, Franz; Stetten, David von; Heck, M.; Sommer, Monika; Hildebrand, Peter W.; Scheerer, Patrick

doi:10.1038/ncomms5801

Cited by 150 publications

(141 citation statements)

References 60 publications

Supporting

Mentioning

125

Contrasting

Order By: Relevance

“…Due to proton uptake in R*H + , the R3.50 side chain is liberated from the intrahelical salt-bridge to E3.49 and forms a new hydrogen bond to Y5.58 (16,42). This finding is in agreement with the crystal structure of the R*H + ·ArrFL-1 complex, which shows this interaction and also the free η-nitrogen of R3.50 forming a strong hydrogen bond to the arrestin finger loop ( Figure 1B) (17). The strongest band in the PBS of R*H + ·ArrFL-1 is the positive band at 1657 cm -1 , whose intensity increases in 2 H 2 O, probably due to the deuteration-induced shift of the negative arginine band at 1662 cm -1 .…”

Section: Arrfl-1/ph Dual Titration Assay -Insupporting

confidence: 80%

“…However, the interaction between R3.50 and the finger loop is present in the crystal structure of native lightactivated rhodopsin in complex with the arrestin finger loop peptide ArrFL-1 (Figure1B). In this crystal structure, distances are such that two strong intermolecular hydrogen bonds between R3.50-G75 1 (2.7 Å) and the backbones of K8.48-G76 1 (3.0 Å) are formed, and also an additional intramolecular R3.50-Y5.58 (3.2 Å) bond is within range of hydrogen bonding (17).…”

mentioning

confidence: 99%

“…Therefore, to allow more straightforward and unambiguous interpretation of our data, we made use of ArrFL-1, a synthetic peptide derived from the finger loop of arrestin-1 (rod visual arrestin, 67 YGQEDIDVMGL 77 ) (31). The finger loop constitutes the key interaction site of arrestins with their cognate GPCRs (32) and ArrFL-1 has been shown to compete with full-length arrestin for the same receptor binding site (17). This makes ArrFL-1 a robust and powerful tool to investigate binding of the arrestin finger loop to rhodopsin.…”

mentioning

confidence: 99%

See 2 more Smart Citations

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Elgeti

Kazmin

Rose

et al. 2018

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Signaling of the prototypical G protein coupled receptor (GPCR) rhodopsin through its cognate G protein transducin (G t ) is quenched when arrestin binds to the activated receptor. Although the overall architecture of the rhodopsin-arrestin complex is known, many questions regarding its specificity remain unresolved. Here, using FTIR difference spectroscopy and a dual pH/ peptide titration assay, we show that rhodopsin maintains certain flexibility upon binding the "finger loop" of visual arrestin (prepared as synthetic peptide ArrFL-1). We found that two distinct complexes can be stabilized depending on the protonation state of E3.49 in the conserved (D)ERY motif. Both complexes exhibit different interaction modes and affinities of ArrFL-1 binding. The plasticity of the receptor within the rhodopsin/ArrFL-1 complex stands in contrast to the complex with the C-terminus of the G t α-subunit (GαCT), which stabilizes only one specific substate out of the conformational ensemble. However, GαCT binding and both ArrFL-1 binding modes involve a direct interaction to conserved R3.50, as determined by site-directed mutagenesis. Our findings highlight the importance of receptor conformational flexibility and cytoplasmic proton uptake for modulation of rhodopsin signaling and thereby extend the picture provided by crystal structures of the rhodopsin/arrestin and rhodopsin/ArrFL-1 complexes. Furthermore, the two binding modes of ArrFL-1 identified here involve motifs of conserved amino acids, which indicates that our results may have elucidated a common modulation mechanism of class A GPCR-G protein/-arrestin signaling.Unlike other class A GPCRs the photoreceptor rhodopsin contains the light-sensitive cofactor retinal as a ligand, which is covalently bound to the opsin apoprotein. Upon light-induced cis/trans isomerization of retinal and subsequent alterations within the binding pocket, the signal propagates via GPCR-conserved interhelical networks towards the cytoplasmic surface of the receptor, where major structural rearrangements take place. The most prominent structural change is the rotational outward tilt of transmembrane (TM) helix 6, which opens a cytoplasmic binding crevice and thus constitutes the main prerequisite for binding and activation of signaling via the G protein transducin (1, 2). Even though first identified in rhodopsin, for many other GPCRs homologous structural changes have been shown to exist, and a common structural and functional framework for GPCRs is being developed (3).

show abstract

Section: Arrfl-1/ph Dual Titration Assay -Insupporting

confidence: 80%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Elgeti

Kazmin

Rose

et al. 2018

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…We focused our mutagenesis approach on the FLR of βarr1 because this region mediates an essential interaction with the receptor transmembrane core (13,15) that stabilizes the GPCRβarr complex core conformation (16). Disrupting this interaction through βarr1 mutagenesis, we reasoned, would allow us to obtain a βarr1 that predominantly forms GPCR-βarr tail conformation complexes, and not any core-conformation complexes, when bound to GPCRs.…”

Section: Resultsmentioning

confidence: 99%

Distinct conformations of GPCR–β-arrestin complexes mediate desensitization, signaling, and endocytosis

Cahill

Thomsen

Tarrasch

et al. 2017

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

303

308

View full text Add to dashboard Cite

β-Arrestins (βarrs) interact with G protein-coupled receptors (GPCRs) to desensitize G protein signaling, to initiate signaling on their own, and to mediate receptor endocytosis. Prior structural studies have revealed two unique conformations of GPCR-βarr complexes: the "tail" conformation, with βarr primarily coupled to the phosphorylated GPCR C-terminal tail, and the "core" conformation, where, in addition to the phosphorylated C-terminal tail, βarr is further engaged with the receptor transmembrane core. However, the relationship of these distinct conformations to the various functions of βarrs is unknown. Here, we created a mutant form of βarr lacking the "finger-loop" region, which is unable to form the core conformation but retains the ability to form the tail conformation. We find that the tail conformation preserves the ability to mediate receptor internalization and βarr signaling but not desensitization of G protein signaling. Thus, the two GPCR-βarr conformations can carry out distinct functions.O ver the past decade, significant efforts have been made to understand the molecular properties and regulatory mechanisms that control the function of β-arrestin (βarr) interactions with G protein-coupled receptors (GPCRs) (1, 2). Once activated, GPCRs initiate a highly conserved signaling and regulatory cascade marked by interactions with: (i) heterotrimeric G proteins, which mediate their actions largely by promoting second-messenger generation (3); (ii) GPCR kinases (GRKs), which phosphorylate activated conformations of receptors (4); and (iii) βarrs, which bind to the phosphorylated receptors to mediate desensitization of G protein signaling and receptor internalization (5, 6). In addition to their canonical function of desensitization and internalization, βarrs have been appreciated as independent signaling units by virtue of their crucial role as both adaptors and scaffolds for an increasing number of signaling pathways (7-11).There are two driving forces that mediate βarr interactions with an activated GPCR: phosphorylation of the C-terminal tail of the receptor by GRKs and/or binding to the transmembrane core of the receptor. How each of these interactions contributes to βarr functionality remains unclear. Moreover, GPCRs tend to either interact with βarr transiently, termed "class A" GPCRs [e.g., β 2 -adrenergic receptor (β 2 AR)], or tightly, known as "class B" GPCRs [e.g., vasopressin type 2 receptor (V 2 R)]. For the current study, we use a previously described chimeric β 2 V 2 R construct, which comprises the β 2 AR with its C-terminal tail exchanged with the V 2 R C-terminal tail (12-14). The β 2 V 2 R construct provides an ideal system for studying a GPCR-βarr complex in vitro, because it maintains identical pharmacological properties to the WT β 2 AR and has a robustly increased class B affinity for βarr1, which allows stable β 2 V 2 R-βarr complexes to be formed and purified.Structural insights have shed some light onto the complexity of the interaction between GPCRs and βarrs. A recent struc...

show abstract

“…Recently, visualization of the GPCR-arrestin interface was achieved by serial femtosecond X-ray laser crystallography of a rhodopsin/arrestin complex (24). Additional insight was gained through cocrystallization studies of an arrestin finger loop peptide and rhodopsin (25), and by electron microscopy and deuterium exchange analysis of a β-arrestin1 complex with a β 2 AR-vasopressin 2 receptor C-terminal tail fusion (26) (25). This conformational stabilization is similar to that induced by G s interaction with the β 2 AR (27).…”

Section: Icl1-9mentioning

confidence: 99%

β-arrestin–biased signaling through the β ₂ -adrenergic receptor promotes cardiomyocyte contraction

Carr

Schilling

Song

et al. 2016

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

102

View full text Add to dashboard Cite

β-adrenergic receptors (βARs) are critical regulators of acute cardiovascular physiology. In response to elevated catecholamine stimulation during development of congestive heart failure (CHF), chronic activation of G s -dependent β 1 AR and G i -dependent β 2 AR pathways leads to enhanced cardiomyocyte death, reduced β 1 AR expression, and decreased inotropic reserve. β-blockers act to block excessive catecholamine stimulation of βARs to decrease cellular apoptotic signaling and normalize β 1 AR expression and inotropy. Whereas these actions reduce cardiac remodeling and mortality outcomes, the effects are not sustained. Converse to G-protein-dependent signaling, β-arrestin-dependent signaling promotes cardiomyocyte survival. Given that β 2 AR expression is unaltered in CHF, a β-arrestin-biased agonist that operates through the β 2 AR represents a potentially useful therapeutic approach. Carvedilol, a currently prescribed nonselective β-blocker, has been classified as a β-arrestin-biased agonist that can inhibit basal signaling from βARs and also stimulate cell survival signaling pathways. To understand the relative contribution of β-arrestin bias to the efficacy of select β-blockers, a specific β-arrestin-biased pepducin for the β 2 AR, intracellular loop (ICL)1-9, was used to decouple β-arrestin-biased signaling from occupation of the orthosteric ligand-binding pocket. With similar efficacy to carvedilol, ICL1-9 was able to promote β 2 AR phosphorylation, β-arrestin recruitment, β 2 AR internalization, and β-arrestin-biased signaling. Interestingly, ICL1-9 was also able to induce β 2 AR-and β-arrestin-dependent and Ca 2+ -independent contractility in primary adult murine cardiomyocytes, whereas carvedilol had no efficacy. Thus, ICL1-9 is an effective tool to access a pharmacological profile stimulating cardioprotective signaling and inotropic effects through the β 2 AR and serves as a model for the next generation of cardiovascular drug development.B eta-antagonists, also known as β-blockers, have been indicated for the treatment of pathological cardiac diseases, including congestive heart failure (CHF) and high blood pressure, for decades (1, 2). A select number of these agents, including the clinically used carvedilol, have been identified as β-arrestinbiased agonists of β-adrenergic receptors based on their ability to promote β-arrestin-dependent signaling over G-protein activation (3, 4). It is believed that the β-arrestin activation may provide additional cardioprotection based on its ability to mediate antiapoptotic signaling. As these are orthosteric ligands, there have been no means to decouple the activation of receptor-dependent β-arrestin signaling from the occupation of the orthosteric ligandbinding pocket to study their independent contribution to its efficacy as these properties appear inherently linked.Recently, we described the characterization of a library of modulators of the β 2 -adrenergic receptor (β 2 AR) known as pepducins (5). Pepducins are lipidated peptides derived from the intracel...

show abstract

Crystal structure of a common GPCR-binding interface for G protein and arrestin

Cited by 150 publications

References 60 publications

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Distinct conformations of GPCR–β-arrestin complexes mediate desensitization, signaling, and endocytosis

β-arrestin–biased signaling through the β ₂ -adrenergic receptor promotes cardiomyocyte contraction

Contact Info

Product

Resources

About

Crystal structure of a common GPCR-binding interface for G protein and arrestin

Cited by 150 publications

References 60 publications

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Distinct conformations of GPCR–β-arrestin complexes mediate desensitization, signaling, and endocytosis

β-arrestin–biased signaling through the β 2 -adrenergic receptor promotes cardiomyocyte contraction

Contact Info

Product

Resources

About

β-arrestin–biased signaling through the β ₂ -adrenergic receptor promotes cardiomyocyte contraction