Structure-activity studies of the thrombin receptor activating peptide

Sabo, Tamar; Genevieve, David; Motola, L.; Brodt, Pnina; Barak, Ruth; Elhanaty, E.

doi:10.1016/0006-291x(92)91099-c

Cited by 58 publications

(45 citation statements)

References 10 publications

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“…Structurefunction studies in terms of platelet activation have been conducted on numerous SFLLRN-based PAR-1 agonist peptides (9,(28)(29)(30)(31)(32)(33)(34). Generally speaking, the peptides require at least the N-terminal pentapeptide with a free amino group at position 1, a certain aromatic residue such as phenylalanine at position 2, and a basic residue such as arginine at position 5.…”

Section: Resultsmentioning

confidence: 99%

Design, synthesis, and biological characterization of a peptide-mimetic antagonist for a tethered-ligand receptor

Andrade‐Gordon¹,

Maryanoff²,

Derian³

et al. 1999

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

179

127

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Protease-activated receptors (PARs) represent a unique family of seven-transmembrane G protein-coupled receptors, which are enzymatically cleaved to expose a truncated extracellular N terminus that acts as a tethered activating ligand. PAR-1 is cleaved and activated by the serine protease ␣-thrombin, is expressed in various tissues (e.g., platelets and vascular cells), and is involved in cellular responses associated with hemostasis, proliferation, and tissue injury. We have discovered a series of potent peptide-mimetic antagonists of PAR-1, exemplified by RWJ-56110. Spatial relationships between important functional groups of the PAR-1 agonist peptide epitope SFLLRN were employed to design and synthesize candidate ligands with appropriate groups attached to a rigid molecular scaffold. Prototype RWJ-53052 was identified and optimized via solid-phase parallel synthesis of chemical libraries. RWJ-56110 emerged as a potent, selective PAR-1 antagonist, devoid of PAR-1 agonist and thrombin inhibitory activity. It binds to PAR-1, interferes with PAR-1 calcium mobilization and cellular function (platelet aggregation; cell proliferation), and has no effect on PAR-2, PAR-3, or PAR-4. By flow cytometry, RWJ-56110 was confirmed as a direct inhibitor of PAR-1 activation and internalization, without affecting N-terminal cleavage. At high concentrations of ␣-thrombin, RWJ-56110 fully blocked activation responses in human vascular cells, albeit not in human platelets; whereas, at high concentrations of SFLLRN-NH 2, RWJ-56110 blocked activation responses in both cell types. Thus, thrombin activates human platelets independently of PAR-1, i.e., through PAR-4, which we confirmed by PCR analysis. Selective PAR-1 antagonists, such as RWJ-56110, should serve as useful tools to study PARs and may have therapeutic potential for treating thrombosis and restenosis.

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Section: Resultsmentioning

confidence: 99%

Design, synthesis, and biological characterization of a peptide-mimetic antagonist for a tethered-ligand receptor

Andrade‐Gordon¹,

Maryanoff²,

Derian³

et al. 1999

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

179

127

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“…Platelets were suspended in the same buffer at 3.5 ϫ 10 8 platelets/ml. This suspension (15 ml) was treated with control saline buffer or with 50 M thrombin receptor-activating peptide (TRAP) (27)(28)(29) at 37°C for 5 min. Platelets were pelleted, and supernatants were collected and concentrated to dryness.…”

Section: Methodsmentioning

confidence: 99%

The Hippocampal Cholinergic Neurostimulating Peptide, the N-terminal Fragment of the Secreted Phosphatidylethanolamine-binding Protein, Possesses a New Biological Activity on Cardiac Physiology

Goumon

Angelone

Schoentgen

et al. 2004

Journal of Biological Chemistry

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Phosphatidylethanolamine-binding protein (PEBP), alternatively named Raf-1 kinase inhibitor protein, is the precursor of the hippocampal cholinergic neurostimulating peptide (HCNP) corresponding to its natural N-terminal fragment, previously described to be released by hippocampal neurons. PEBP is a soluble cytoplasmic protein, also associated with plasma and reticulum membranes of numerous cell types. In the present report, using biochemistry and cell biology techniques, we report for the first time the presence of PEBP in bovine chromaffin cell, a well described secretion model. We have examined its presence at the subcellular level and characterized this protein on both secretory granule membranes and intragranular matrix. In addition, its presence in bovine chromaffin cell and platelet exocytotic medium, as well as in serum, was reported showing that it is secreted. Like many other proteins that lack signal sequence, PEBP may be secreted through non-classic signal secretory mechanisms, which could be due to interactions with granule membrane lipids and lipid rafts. By two-dimensional liquid chromatography-tandem mass spectrometry, HCNP was detected among the intragranular matrix components. The observation that PEBP and HCNP were secreted with catecholamines into the circulation prompted us to investigate endocrine effects of this peptide on cardiovascular system. By using as bioassay an isolated and perfused frog (Rana esculenta) heart preparation, we show here that HCNP acts on the cardiac mechanical performance exerting a negative inotropism and counteracting the adrenergic stimulation of isoproterenol. All together, these data suggest that PEBP and HCNP might be considered as new endocrine factors involved in cardiac physiology.Chromaffin cells are derived from the neural crest and constitute the adrenal medulla. In the adrenal medullary chromaffin cell, secretory granules contain a complex mixture of proteins and peptides that are co-released with catecholamines into the circulation in response to splanchnic nerve stimulation (1) . Among the high molecular mass water-soluble proteins, proenkephalin-A and the chromogranins family constitute the major constituents of chromaffin granules with other neuropeptides (neuropeptide Y, vasointestinal peptide, and others). These proteins are actively processed in the intragranular matrix to peptides with various molecular weight (1-3). Recently, using highly sensitive proteomic techniques we have characterized the maturation products of proenkephalin-A (4) and established the presence of other unexpected proteins (5).Phosphatidylethanolamine-binding protein (PEBP) 1 (6, 7)/ Raf-1 kinase inhibitor protein (8) has previously been described in brain and adrenal gland (9, 10). PEBP is a 21-kDa protein initially described as a cytoplasmic protein and later found to be associated with plasma or reticulum membranes (11). This protein has been found in numerous tissues of many species, including rat testis, liver, kidney, and human platelets (6, 9, 10), indicating its ubi...

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“…The cleavage unmasks a new amino terminus, which acts as a tethered ligand that promotes receptor activation. A synthetic peptide consisting o f as few as Five amino acids o f the new amino terminus has been shown to fully activate the thrombin receptor [7][8][9].…”

Section: Introductionmentioning

confidence: 99%

Vascular Effects of Proteinase-Activated Receptor 2 Agonist Peptide

Emilsson

Wahlestedt²,

Sun

et al. 1997

J Vasc Res

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Proteinase-activated receptor 2 (PAR-2) is a G protein-coupled receptor related to the thrombin receptor. PAR-2 can be activated by trypsin and by synthetic peptides corresponding to the new amino terminus generated by activating proteolytic cleavage. We show in this report that intravenous injection of PAR-2 agonist peptides has dramatic effects on arterial blood pressure in anesthetized rats. The peptide SLIGRLETQPPI, at 150 nmol/kg, transiently decreased the mean arterial pressure from 104 to 60 mm Hg. The hypotensive response was dose-dependent, and was not secondary to effects on central vasoregulatory systems, heart rate, or the kidneys. A nitric oxide synthase inhibitor attenuated the hypotensive response induced by the PAR-2 agonist peptide. Further experiments in vitro, on preparations of rat femoral artery and vein, showed that PAR-2 agonist peptide elicited a dose-dependent relaxation of both types of vessel. Removal of the endothelium abolished the agonist peptide-induced relaxation. Our results demonstrate that activation of PAR-2 can modulate vascular tone, and that this response was an effect mediated at least partly by nitric oxide. The effect on blood vessels further suggests that the physiological activator of this proteolytically activated receptor is an enzyme present and active in the blood, possibly after a vascular injury.

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Structure-activity studies of the thrombin receptor activating peptide

Cited by 58 publications

References 10 publications

Design, synthesis, and biological characterization of a peptide-mimetic antagonist for a tethered-ligand receptor

Design, synthesis, and biological characterization of a peptide-mimetic antagonist for a tethered-ligand receptor

The Hippocampal Cholinergic Neurostimulating Peptide, the N-terminal Fragment of the Secreted Phosphatidylethanolamine-binding Protein, Possesses a New Biological Activity on Cardiac Physiology

Vascular Effects of Proteinase-Activated Receptor 2 Agonist Peptide

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