Related Contribution of Specific Helix 2 and 7 Residues to Conformational Activation of the Serotonin 5-HT2A Receptor

Sealfon, Stuart C.; Chi, Ling; Ebersole, Barbara J.; Rodic, Vladimir; Zhang, Daqun; Ballesteros, Juan A.; Weinstein, Harel

doi:10.1074/jbc.270.28.16683

Cited by 207 publications

(217 citation statements)

References 24 publications

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“…Several works have suggested a Asp-2.50/Asn-7.49 direct interaction in the active state of GPCRs (36,37). These two residues are found interchanged in the gonadotropin-releasing hormone receptor and engineering such a swap leads to functional receptors in a number of cases (36,37).…”

Section: Resultsmentioning

confidence: 99%

An Activation Switch in the Rhodopsin Family of G Protein-coupled Receptors

Urizar

Claeysen

Deupí

et al. 2005

Journal of Biological Chemistry

107

100

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We aimed at understanding molecular events involved in the activation of a member of the G protein-coupled receptor family, the thyrotropin receptor. We have focused on the transmembrane region and in particular on a network of polar interactions between highly conserved residues. Using molecular dynamics simulations and site-directed mutagenesis techniques we have identified residue Asn-7.49, of the NPxxY motif of TM 7, as a molecular switch in the mechanism of thyrotropin receptor (TSHr) activation. Asn-7.49 appears to adopt two different conformations in the inactive and active states. These two states are characterized by specific interactions between this Asn and polar residues in the transmembrane domain. The inactive gauche؉ conformation is maintained by interactions with residues Thr-6.43 and Asp-6.44. Mutation of these residues into Ala increases the constitutive activity of the receptor by factors of ϳ14 and ϳ10 relative to wild type TSHr, respectively. Upon receptor activation Asn-7.49 adopts the trans conformation to interact with Asp-2.50 and a putatively charged residue that remains to be identified. In addition, the conserved Leu-2.46 of the (N/S)LxxxD motif also plays a significant role in restraining the receptor in the inactive state because the L2.46A mutation increases constitutive activity by a factor of ϳ13 relative to wild type TSHr. As residues Leu-2.46, Asp-2.50, and Asn-7.49 are strongly conserved, this molecular mechanism of TSHr activation can be extended to other members of the rhodopsin-like family of G protein-coupled receptors.Genome sequencing projects have identified the G proteincoupled receptor (GPCR) 1 family as one of the largest class of proteins with more than 800 human sequences. These sequences have been classified into five main families, glutamate, rhodopsin, adhesion, frizzled/taste2, and secretin (1). GPCRs are involved in passing chemical signals across the cell membrane. The incoming signal may arrive in the form of neurotransmitters, peptides, divalent cations, proteases, hormones, and sensory stimuli such as photons and gustatory or odorant molecules (2, 3). Atomic level details of a three-dimensional structure of a GPCR are only known for the inactive form of rhodopsin, the light photoreceptor protein of rod cells (4, 5). Rhodopsin is formed by an extracellular N terminus, seven ␣-helices, which cross the cellular membrane (TM 1-TM 7) connected by hydrophilic loops, and a cytoplasmic C terminus containing an ␣-helix (HX 8) parallel to the cell membrane. The overall structure of the helical bundle seems common in the rhodopsin-like GPCR family because of the large number of conserved sequence patterns in these 7 TM segments (6): GN in TM 1, (N/S)LxxxD in TM 2, (D/E)RY in TM 3, CWxP in TM 6, and NPxxY(x) 5,6 F in TM 7 and HX 8, among others. Activation of GPCRs is commonly discussed in terms of the extended ternary complex model, which proposes an equilibrium between inactive and active states (7). Recent advances in the ␤ 2 -adrenergic receptor have shown that ag...

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Section: Resultsmentioning

confidence: 99%

An Activation Switch in the Rhodopsin Family of G Protein-coupled Receptors

Urizar

Claeysen

Deupí

et al. 2005

Journal of Biological Chemistry

107

100

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“…In the case of the RH-based modeling, constraints resulting from the alignment of the 5-HT receptor sequences to the template led to an electrostatic interaction between Asp 2.50(h100,m99) and Asn 7.49(h376,m375) . This interaction was already evidenced between Asp 2.50(120) and Asn 7.49(396) in the human 5HT 2A R (28). With the BR-based models, the orientation of TMD7 with respect to TMD2 had to be slightly modified to establish a similar interaction between Asp 2.50(h100,m99) and Asn 7.49(h376,m375) .…”

Section: Computer Modeling Of Human and Mouse Wild-type 5-ht 2bmentioning

confidence: 88%

The Serotonin Binding Site of Human and Murine 5-HT2BReceptors

Manivet¹,

Schneider²,

Smith³

et al. 2002

Journal of Biological Chemistry

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Bacteriorhodopsin and rhodopsin crystal structures were used as templates to build structural models of the mouse and human serotonin (5-HT)-2B receptors (5-HT 2B Rs). Serotonin was docked to the receptors, and the amino acids predicted to participate to its binding were subjected to mutagenesis. 5-HT binding affinity and 5-HT-induced inositol triphosphate production were measured in LMTK ؊ cells transfected with either wildtype or mutated receptor genes. According to these measurements, the bacteriorhodopsin-based models of the 5-HT 2B Rs appear more confident than the rhodopsin-based ones. Residues belonging to the transmembrane domains 3 and 6, i.e. Of the 14 mammalian serotonin (5-hydroxytryptamine (5-HT) 1 ) receptor subtypes, all but one (5-HT 3 ) belong to the super-family of G-protein-coupled receptors (GPCRs) (1). The 5-HT 2 subtype comprises three closely related receptors, which are 5-HT 2A , 5-HT 2B , and 5-HT 2C . Amino acid sequences of the rat (2), mouse (3), and human (4) 5-HT 2B receptors (5-HT 2B Rs) are highly similar at the level of the predicted transmembrane domains (TMDs); they exhibit 88, 82, and 79% homology upon comparison of mouse and rat, human and mouse, and human and rat receptors, respectively (5). Moreover, significant correlations were established between the pharmacological profiles of human and rat (or human and mouse) 5-HT 2B Rs but not between those of mouse and rat (5). Nevertheless, some compounds (e.g. certain ergolines and benzoylpiperidines) can be used to discriminate pharmacologically between human and rat 5-HT 2B Rs (6).Structural descriptions of these receptors and of their binding interactions are required to rationalize the above findings. In the absence of any 5-HT receptor three-dimensional structure, computer modeling studies were undertaken in the present study. Our aim was to explain the different 5-HT pK D values measured for the rat, mouse, and human 5-HT 2B Rs (7.5, 5.8, and 7.9, respectively) and to better understand the mechanism of 5-HT 2B R activation.Two main strategies have already been used to build GPCR three-dimensional models. The first one relied on the use of the structure of bacteriorhodopsin (BR) at a 3-Å resolution as a homology template (7,8). The second strategy, already used for three-dimensional-modeling of the 5-HT 2A (9, 10) and 5-HT 2C receptors (10, 11), aimed at constructing models de novo by only using the ϳ9-Å structure of bovine rhodopsin (RH) as a guide to orient the TMDs. These studies were the starting point in the elucidation of the conformational switch between the inactive and active states of GPCRs. A key feature of these approaches involved the replacement of the network of constraints in the inactive ground state of the receptor by a ligand-induced set of interactions in the activated state (12)(13)(14).In the present work, the construction of three-dimensional models of the mouse and human 5-HT 2B Rs was carried out using either the high resolution BR structure or the recently described crystal structure of bovine RH (1...

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“…The substitution of the Asn residue in the NPXXY motif in different GPCRs has been shown to affect the activation of adenylyl cyclase [19], phospholipase C [20], [21], phospholipase D [22], and internalization pathways [23 -24].…”

Section: Homology Of Fatty Acid Binding Receptors To Class a Gpcrs Amentioning

confidence: 99%

Fatty Acid Binding Receptors and Their Physiological Role in Type 2 Diabetes

Swaminath

2008

Archiv der Pharmazie

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G‐protein‐coupled receptors (GPCRs) respond to various physiological ligands such as photons, ions, and small molecules that include amines, fatty acids, and amino acids to peptides, proteins and steroids. Therefore, this family of proteins represents an attractive target for biopharmaceutical research [1]. The physiological role of fatty acids and other lipid molecules as important signal mediators is well studied in various metabolic pathways [2]. Acute administration of free fatty acids (FFAs) stimulates insulin release. Conversely, chronic exposure to high levels of free fatty acids leads to impairment of β cell function and lipotoxicity. However, the receptors through which these fatty acids and lipids act were unknown, until the identification of fatty acid binding receptors: GPR40, GPR41, GPR43, and GPR119. Based on their tissue‐expression profile, and pharmacologic analysis, the fatty acid binding receptors along with lipid binding receptor GPR119 are linked to diabetes and obesity. They play a critical role in the metabolic regulation of insulin release and glucose homeostasis. In this review, the mechanism of receptor activation, pharmacology, and the physiological functions of the fatty acid binding receptors will be discussed.

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Related Contribution of Specific Helix 2 and 7 Residues to Conformational Activation of the Serotonin 5-HT2A Receptor

Cited by 207 publications

References 24 publications

An Activation Switch in the Rhodopsin Family of G Protein-coupled Receptors

An Activation Switch in the Rhodopsin Family of G Protein-coupled Receptors

The Serotonin Binding Site of Human and Murine 5-HT2BReceptors

Fatty Acid Binding Receptors and Their Physiological Role in Type 2 Diabetes

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