A Proposed Model of Bradykinin Bound to the Rat B2 Receptor and Its Utility for Drug Design

Kyle, Donald J.; Chakravarty, Sarvajit; Sinsko, Jacqueline A.; Stormann, Thomas M.

doi:10.1021/jm00035a015

Cited by 82 publications

(71 citation statements)

References 9 publications

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“…We conclude from these results that BK, when bound to the B2 receptor, reaches from the extracellular surface of the receptor adjacent to EC-IV, down two helical turns along the interior face of TM-VI, and across into a pocket bordered by the interior face of TM-III, and presumably TM-V and -VI, and adjacent to Ser 111 . These results are in remarkably good agreement with a model of BK bound to the B2 receptor proposed by Kyle based on structural homology Table II and Table III modeling, molecular modeling, and systematic conformational searching methods of BK and the receptor (22). Two-dimensional NMR and molecular modeling of BK in various solvents and micelles have revealed that Ser 6 through Arg 9 adopt a ␤-turn-like structure when in a hydrophobic environment (23,24).…”

Section: Discussionsupporting

confidence: 86%

“…We have previously shown that the N terminus of the bound BK is located within 3 Å of extracellular Cys 20 in EC-I and Cys 277 in EC-IV (15). BK binding is dependent on two Asp residues, Asp 268 and Asp 286 , located in EC-IV (12) and with which either the N terminus or the guanidinium side chain of Arg 1 is believed to interact (22). The binding of BK also requires Phe 259 and Thr 263 located two and one helical turns, respectively, below EC-IV on the interior face in TM-VI (11,13,14).…”

Section: Discussionmentioning

confidence: 99%

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A Single Position in the Third Transmembrane Domains of the Human B1 and B2 Bradykinin Receptors Is Adjacent to and Discriminates between the C-terminal Residues of Subtype-selective Ligands

Fathy

Mathis

Leeb³

et al. 1998

Journal of Biological Chemistry

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In order to identify agonist-and antagonist-binding epitopes in the human B1 and B2 bradykinin (BK) receptors, we exploited the ability of these receptors to discriminate between peptide ligands that differ only by the absence (B1) and presence (B2) of a C-terminal Arg. This was done by constructing chimeric proteins in which specific domains were exchanged between these receptors as recently described by us (Leeb, T., Mathis, S. A., and LeebLundberg, L. M. Seven-transmembrane domain GPCR 1 constitute by far the largest family of plasma membrane receptors. These receptors bind ligands of widely diverse origins, and are unsurpassed as therapeutic targets. Consequently, much effort has been devoted to mapping of the binding sites for agonist and antagonist ligands in these receptors (1, 2). Even though peptides are the most common class of ligands for GPCR, few peptide GPCR have been investigated thus far, and in most of those cases the identity of the peptide-binding epitopes remains elusive.Receptors for kinins, pro-inflammatory peptides 8 -10 amino acids in length, have been classified into two subtypes, termed B1 and B2 (3), and are members of the GPCR superfamily (4, 5). These receptor subtypes, although only 36% identical, discriminate between peptide agonists that differ only in their C-terminal residue; BK binds to the B2 receptor, whereas the C-terminally truncated carboxypeptidase fragments desArg 9 -BK and des-Arg 10 -Lys-BK, or des-Arg 10 -KD, bind to the B1 receptor. Several high affinity B2 receptor-selective decapeptide antagonists structurally derived from BK have been developed, including NPC17731 and HOE140 (6 -8). Interestingly, the fact that the des-Arg 10 analogs of these peptides act as high affinity B1 receptor-selective antagonists emphasizes the significance of the C-terminal Arg in receptor subtype selectivity (9 -11).In the B2 receptor, extensive analysis of most of the TMs and a significant amount of the ECs by alanine-scanning mutagenesis has yielded no information about residues important for antagonist binding and has identified only a few residues important for agonist binding (12-15). We recently developed a novel, potentially more effective strategy for mapping the binding sites in kinin receptors, which is based on the identification of receptor epitopes that enable these receptors to discriminate between ligands (11). This strategy involves the exchange of individual TMs between the B1 and B2 receptor subtypes and the subsequent exchange of non-conserved residues that are possible candidates for discriminatory action. This approach is intrinsically more reliable than alanine-scanning mutagenesis as it yields in sequence both loss-of-function and gain-of-function mutations.In an initial study, we used this strategy to identify specific residues in TM-VI of the human WT B1 and B2 receptors that are partially responsible for enabling these receptor subtypes to discriminate between peptide agonists (11). In the present study, we analyzed the role of TM-III in peptide ligand discrimi...

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Section: Discussionsupporting

confidence: 86%

Section: Discussionmentioning

confidence: 99%

A Single Position in the Third Transmembrane Domains of the Human B1 and B2 Bradykinin Receptors Is Adjacent to and Discriminates between the C-terminal Residues of Subtype-selective Ligands

Fathy

Mathis

Leeb³

et al. 1998

Journal of Biological Chemistry

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“…Our models were designed to give ideas for further mutagenesis experiments and possible binding modes of agonists and antagonists. Our models differ somewhat from those previously described (52,53). The major difference centers on each group's choice for the ends of the ␣-helices, particularly TM-6 and TM-7.…”

Section: Discussioncontrasting

confidence: 63%

Mutations in the B2 Bradykinin Receptor Reveal a Different Pattern of Contacts for Peptidic Agonists and Peptidic Antagonists

Jarnagin¹,

Bhakta²,

Zuppan³

et al. 1996

Journal of Biological Chemistry

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The B 2 bradykinin receptor, a seven-helix transmembrane receptor, binds the inflammatory mediator bradykinin (BK) and the structurally related peptide antagonist HOE-140. The binding of HOE-140 and the binding of bradykinin are mutually exclusive and competitive. Fifty-four site-specific receptor mutations were made. BK's affinity is reduced 2200-fold by F261A, 490-fold by T265A, 60-fold by D286A, and 3-10-fold by N200A, D268A, and Q290A. In contrast, HOE-140 affinity is reduced less than 7-fold by F254A, F261A, Y297A, and Q262A. Bradykinin is a central initiator of acute and chronic inflammation and the associated pain and edema. Most of the acute and many of the chronic responses to bradykinin are mediated by B 2 bradykinin receptors (1, 2), while some of the chronic responses to bradykinin are mediated by B 1 bradykinin receptors (3). B 1 and B 2 bradykinin receptors are members of the G-protein-coupled seven-transmembrane (GPC-7TM) 1 receptor superfamily (4 -7). For B 1 receptors, [des-Arg 9 ]bradykinin is a more potent agonist than bradykinin, whereas B 2 receptors bind and respond to bradykinin about 10,000-fold more effectively than [des-Arg 9 ]bradykinin. Several peptidic B 2 bradykinin antagonists have been identified; these compounds reduce pain and inflammation (8 -12). Peptidic antagonists also reduce death from experimental shock (13-15). Bradykinin receptor antagonists are potentially useful in the treatment of pain, acute and chronic inflammation, shock, allergic or infectious rhinitis, and asthma. The peptidic antagonists are useful tools; but, to date, these compounds have made poor human therapeutic agents because of their poor bioavailability and formulation difficulties (16,17). The discovery of a nonpeptide antagonist of bradykinin would improve the prospects of treating bradykinin-instigated inflammation, pain, or edema. Thus, we have focused on a molecular understanding of the bradykinin receptor ligand binding site, believing that this information may help in the discovery and design of nonpeptidic antagonists.We used the results from molecular modeling studies of B 2 bradykinin receptors and NMR studies of peptidic agonists and antagonists to generate a number of models for agonist binding to the B 2 BKR. These models were tested by site-directed mutagenesis of the receptor and by making single amino acid changes in agonist and antagonist peptides. The data reveal a disparity between the way peptidic agonists and antagonists bind to the BKR. We attempt to reconcile the disparities by proposing new models of the BKR-ligand complex. -140 (18, 19). The product was HPLC-purified and then characterized by HPLC and mass spectrometry as to its chemical purity (Ն96%), identity, and specific activity (56.5 Ci/mmol). Media and other cell culture additives were from Life Technologies, Inc. Biochemicals and enzymes were from Boehringer Mannheim. Common reagents were from Sigma. EXPERIMENTAL PROCEDURES Materials MethodsStandard molecular biological and cell culture methods were used except as speci...

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“…An early model of BK bound to the rat B 2 receptor based on structural homology modeling with bacteriorhodopsin, molecular modeling, and systematic conformational searching methods led to the identification of two candidate residues, Asp 268 and Asp 286 (human residues Asp 266 and Asp 284 ) in EL-3 near the extracellular ends of TM-6 and TM-7, respectively, with which BK may interact ( Fig. 3) (Kyle et al, 1994). It was proposed that these aspartates interact electrostatically with the N-terminal amino group, the guanidinyl side chain, or both on Arg 1 in BK, a residue absolutely critical for function (Regoli and Barabé, 1980).…”

Section: Agonist and Antagonist Binding Sites In The Receptorsmentioning

confidence: 99%

International Union of Pharmacology. XLV. Classification of the Kinin Receptor Family: from Molecular Mechanisms to Pathophysiological Consequences

Marceau²,

et al. 2005

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A Proposed Model of Bradykinin Bound to the Rat B2 Receptor and Its Utility for Drug Design

Cited by 82 publications

References 9 publications

A Single Position in the Third Transmembrane Domains of the Human B1 and B2 Bradykinin Receptors Is Adjacent to and Discriminates between the C-terminal Residues of Subtype-selective Ligands

A Single Position in the Third Transmembrane Domains of the Human B1 and B2 Bradykinin Receptors Is Adjacent to and Discriminates between the C-terminal Residues of Subtype-selective Ligands

Mutations in the B2 Bradykinin Receptor Reveal a Different Pattern of Contacts for Peptidic Agonists and Peptidic Antagonists

International Union of Pharmacology. XLV. Classification of the Kinin Receptor Family: from Molecular Mechanisms to Pathophysiological Consequences

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