title: Short peptide constructs mimic agonist sites of AT(1)R and BK receptors.
abstract: Extracellular peptide ligand binding sites, which bind the N-termini of angiotensin II (AngII) and bradykinin (BK) peptides, are located on the N-terminal and extracellular loop 3 regions of the AT(1)R and BKRB(1) or BKRB(2) G-protein-coupled receptors (GPCRs). Here we synthesized peptides P15 and P13 corresponding to these receptor fragments and showed that only constructs in which these peptides were linked by S-S bond, and cyclized by closing the gap between them, could bind agonists. The formation of construct-agonist complexes was revealed by electron paramagnetic resonance spectra and fluorescence measurements of spin labeled biologically active analogs of AngII and BK (Toac(1)-AngII and Toac(0)-BK), where Toac is the amino acid-type paramagnetic and fluorescence quencher 2, 2, 6, 6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid. The inactive derivatives Toac(3)-AngII and Toac(3)-BK were used as controls. The interactions characterized by a significant immobilization of Toac and quenching of fluorescence in complexes between agonists and cyclic constructs were specific for each system of peptide-receptor construct assayed since no crossed reactions or reaction with inactive peptides could be detected. Similarities among AT, BKR, and chemokine receptors were identified, thus resulting in a configuration for AT(1)R and BKRB cyclic constructs based on the structure of the CXCR(4), an α-chemokine GPCR-type receptor.
authors.affiliation: Department of Biophysics, Federal University of Sao Paulo, Sao Paulo CEP 04044-020, Brazil.
authors.forename: Douglas D
authors.initials: DD
authors.lastname: Lopes
authors.forename: Renata F F
authors.initials: RF
authors.lastname: Vieira
authors.forename: Luciana
authors.initials: L
authors.lastname: Malavolta
authors.forename: Erick F
authors.initials: EF
authors.lastname: Poletti
authors.forename: Suma I
authors.initials: SI
authors.lastname: Shimuta
authors.forename: Antonio C M
authors.initials: AC
authors.lastname: Paiva
authors.forename: Shirley
authors.initials: S
authors.lastname: Schreier
authors.forename: Laerte
authors.initials: L
authors.lastname: Oliveira
authors.forename: Clovis R
authors.initials: CR
authors.lastname: Nakaie
mesh_terms: D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000819:agonists* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D001920:Bradykinin / Q000819:agonists* / Q000235:genetics / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D044140:Receptor, Angiotensin, Type 1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018002:Receptors, Bradykinin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D010455:Peptides; D044140:Receptor, Angiotensin, Type 1; D018002:Receptors, Bradykinin; D000804:Angiotensin II; D001920:Bradykinin