Identification of Two Amino Acids of the Human Cholecystokinin-A Receptor That Interact with the N-terminal Moiety of Cholecystokinin

Kennedy, Karen; Gigoux, Véronique; Escrieut, Chantal; Maigret, Bernard; Martinez, Jean; Moröder, Luis; Frehel, Daniel; Gully, Danielle; Vaysse, Nicole; Fourmy, Daniel

doi:10.1074/jbc.272.5.2920

Cited by 67 publications

(116 citation statements)

References 41 publications

(38 reference statements)

Supporting

Mentioning

100

Contrasting

Order By: Relevance

“…In contrast, the latter model proposed by the groups of Miller and Lybrand (5), places the carboxyl terminus of CCK adjacent to Trp 39 in the amino-terminal tail region just outside of transmembrane segment one (in the position of the amino terminus of CCK in the other model (26)). This model places the amino terminus of CCK above the dominant binding groove, between the third extracellular loop and covered by the glycosylated distal amino-terminal tail of the receptor (8).…”

Section: Lifetime Measurements For Fluorescent Ligandsmentioning

confidence: 60%

“…One of these is based entirely on indirect evidence from mutagenesis studies (26 -28), whereas the other has utilized both structure-activity data and extensive experimentally derived constraints of spatial approximation between residues in ligand and receptor as shown in a series of photoaffinity labeling studies (5-8). The former model, proposed by Fourmy and his collaborators (26,27), places the carboxyl terminus of CCK as dipping into the intramembranous confluence of helices, and positions the amino terminus of the peptide adjacent to the receptor amino-terminal tail region just outside of transmembrane segment one. Such ligand placement may make it difficult to accommodate the large and varied fluorophores being utilized in the current study.…”

Section: Lifetime Measurements For Fluorescent Ligandsmentioning

confidence: 99%

See 1 more Smart Citation

Environment and Mobility of a Series of Fluorescent Reporters at the Amino Terminus of Structurally Related Peptide Agonists and Antagonists Bound to the Cholecystokinin Receptor

Harikumar

Pinon

Wessels

et al. 2002

Journal of Biological Chemistry

103

View full text Add to dashboard Cite

Fluorescence is a powerful biophysical tool for the analysis of the structure and dynamics of proteins. Here, we have developed two series of new fluorescent probes of the cholecystokinin (CCK) receptor, representing structurally related peptide agonists and antagonists. Each ligand had one of three distinct fluorophores (Alexa 488 , nitrobenzoxadiazolyl, or acrylodan) incorporated in analogous positions at the amino terminus just outside the hormone's pharmacophore. All of the probes bound to the CCK receptor specifically and with high affinity, and intracellular calcium signaling studies showed the chemically modified peptides to be fully biologically active. Quenching by iodide and measurement of fluorescence spectra, anisotropy, and lifetimes were used to characterize the response of the fluorescence of the probe in the peptide-receptor complex for agonists and antagonists. All three fluorescence indicators provided the same insights into differences in the environment of the same indicator in the analogous position for agonist and antagonist peptides bound to the CCK receptor. Each agonist had its fluorescence quenched more easily and showed lower anisotropy (higher mobility of the probe) and shorter lifetime than the analogous antagonist. Treatment of agonist-occupied receptors with a non-hydrolyzable GTP analogue shifted the receptor into its inactive low affinity state and increased probe fluorescence lifetimes toward values observed with antagonist probes. These data are consistent with a molecular conformational change associated with receptor activation that causes the amino terminus of the ligand (situated above transmembrane segment six) to move away from its somewhat protected environment and toward the aqueous milieu.Cell surface receptors present on essentially every excitable cell of the body are important targets for pharmacotherapy. A detailed understanding of the structure of these molecules and the molecular basis of their activation should contribute to the rational design and refinement of ligands for these receptors. Receptor-bound, environmentally sensitive fluorescent reporters can provide information regarding ligand-binding domains (1). In this work, we utilize this approach to gain insight into agonist-and antagonist-binding domains of the type A cholecystokinin (CCK) 1 receptor, a physiologically important member of the rhodopsin/␤-adrenergic receptor family of guanine nucleotide-binding protein (G protein)-coupled receptors.The superfamily of G protein-coupled receptors represents the largest group of membrane receptors. They are remarkable for the diversity in structure of the natural agonist ligands that can activate them and initiate intracellular signaling cascades. These range in size from small photons and odorants to peptides, proteins, and even large viral particles. Tertiary structure determination by x-ray crystallography has provided the most incisive insight into the structure of superfamily members, which bind small ligands in the intramembranous helical bundle domain ...

show abstract

Section: Lifetime Measurements For Fluorescent Ligandsmentioning

confidence: 60%

Section: Lifetime Measurements For Fluorescent Ligandsmentioning

confidence: 99%

Environment and Mobility of a Series of Fluorescent Reporters at the Amino Terminus of Structurally Related Peptide Agonists and Antagonists Bound to the Cholecystokinin Receptor

Harikumar

Pinon

Wessels

et al. 2002

Journal of Biological Chemistry

103

View full text Add to dashboard Cite

show abstract

“…The final model respects most transmembrane arrangements found in the recent x-ray structure of rhodopsin (23). For docking of CCK ligand into the CCK1R binding site, experimental data that identified contact points between residues Trp-39 and Gln-40 and the N-terminal moiety of CCK served as a first constraint to place CCK within the CCK1R grove (14). In a first step, manual docking was achieved by taking into account molecular electrostatic potentials at the top of the receptor grove.…”

Section: Methodsmentioning

confidence: 99%

“…Amino acids within three regions of the CCK1R were identified as belonging to the binding site for CCK. Trp-39 and Gln-40, located at the top of transmembrane helix I, were shown to interact directly with the N-terminal portion of CCK (14). , located in the second extracellular loop, were then shown to interact with the sulfated tyrosine (15,16).…”

Section: Cholecystokinin (Cck)mentioning

confidence: 99%

The Biologically Crucial C Terminus of Cholecystokinin and the Non-peptide Agonist SR-146,131 Share a Common Binding Site in the Human CCK1 Receptor

Escrieut

Gigoux

Archer

et al. 2002

Journal of Biological Chemistry

Self Cite

105

View full text Add to dashboard Cite

The cholecystokinin (CCK) receptor-1 (CCK1R) is a G protein-coupled receptor, which mediates important central and peripheral cholecystokinin actions. Our aim was to progress in mapping of the CCK1R binding site by identifying residues that interact with the methionine and phenylalanine residues of the C-terminal moiety of CCK because these are crucial for its binding and These new and important insights will serve to better understand the activation process of CCK1R and to design or optimize ligands.

show abstract

“…This model is distinct from another extensively-discussed model in the literature that has been based exclusively on receptor mutagenesis studies (Kennedy et al 1997;Gigoux et al 1999). It is important to incorporate as many different types of studies and as many independently-derived constraints as possible in developing such a model.…”

Section: Model Of Agonist-occupied Cck Receptormentioning

confidence: 99%

Molecular Basis of Agonist Binding to the Type A Cholecystokinin Receptor

Miller

Lybrand

2002

Pharmacology & Toxicology

View full text Add to dashboard Cite

Abstract:The receptors for cholecystokinin (CCK) peptides are guanine nucleotide-binding protein-coupled receptors in the rhodopsin/b-adrenergic receptor family. The molecular basis of natural ligand binding to the type A CCK receptor has been studied using ligand structure-activity series, receptor mutagenesis, and photoaffinity labeling studies. These have focused attention on the extracellular loop and tail domains, with the most direct insights coming from intrinsic photoaffinity labeling studies. A model of the binding of CCK to this receptor is consistent with all these studies. This model places the carboxyl terminus of CCK adjacent to the amino-terminal tail outside of transmembrane segment 1, the midregion of the peptide adjacent to the third extracellular loop outside of transmembrane segment 7, and includes a chargecharge interaction between peptide residue tyrosine-sulfate 27 and the arginine residue in the second extracellular loop of the receptor in position 197.Understanding of the molecular basis of agonist binding to receptors and their activation is of great interest and importance for the rational design of receptor-active drugs. The type A cholecystokinin (CCK) receptor is a valid drug target, with particular relevance for agonist drugs that might induce satiety and that could have a role in the pharmacotherapy of obesity. Indeed, orally active small molecule drug candidates with agonist activity at this receptor are being developed (Aquino et al. 1996). A more detailed understanding of the conformation of the active complex of CCK receptor occupied by the natural full agonist ligand, CCK, would help to refine such candidates and to develop new candidate drugs.The CCK receptor is a member of the Class I rhodopsin/ b-adrenergic family of guanine nucleotide-binding protein (G protein)-coupled receptors. This molecule was first identified by affinity labeling using a CCK-like probe (Pearson & Miller 1987) and was later purified, partially sequenced, and had its cDNA cloned (Wank et al. 1992). This assignment to this receptor family is based on sequence homology primarily within the transmembrane domains of member receptors. Being a member of the Class I family provides a particular advantage for the CCK receptor, based on the recent successful solution of a high resolution crystal structure of rhodopsin (Palczewski et al. 2000). It is quite likely that there will be substantial similarity between the conformation of transmembrane helices and bundle formation within the membrane of the CCK receptor and rhodopsin. However, in a recent report, it was demonstrated that the loop and tail positions in rhodopsin are not rel- evant to analogous portions of the CCK receptor (Ding et al. 2002). Homology modeling of the CCK receptor for these regions of rhodopsin yield no binding cleft of adequate size and reasonable position to accommodate the natural peptide ligand. Ligand structure-activity seriesThree types of studies can be particularly useful for understanding the conformation of the CCK receptor and...

show abstract

Identification of Two Amino Acids of the Human Cholecystokinin-A Receptor That Interact with the N-terminal Moiety of Cholecystokinin

Cited by 67 publications

References 41 publications

Environment and Mobility of a Series of Fluorescent Reporters at the Amino Terminus of Structurally Related Peptide Agonists and Antagonists Bound to the Cholecystokinin Receptor

Environment and Mobility of a Series of Fluorescent Reporters at the Amino Terminus of Structurally Related Peptide Agonists and Antagonists Bound to the Cholecystokinin Receptor

The Biologically Crucial C Terminus of Cholecystokinin and the Non-peptide Agonist SR-146,131 Share a Common Binding Site in the Human CCK1 Receptor

Molecular Basis of Agonist Binding to the Type A Cholecystokinin Receptor

Contact Info

Product

Resources

About