title: The snake venom peptide Bj-PRO-7a is a M1 muscarinic acetylcholine receptor agonist.
abstract: Proline-rich peptides from Bothrops jararaca venom (Bj-PRO) were characterized based on the capability to inhibit the somatic angiotensin-converting enzyme. The pharmacological action of these peptides resulted in the development of Captopril, one of the best examples of a target-driven drug discovery for treatment of hypertension. However, biochemical and biological properties of Bj-PROs were not completely elucidated yet, and many recent studies have suggested that their activity relies on angiotensin-converting enzyme-independent mechanisms. Here, we show that Bj-PRO-7a (<EDGPIPP) specifically activates [Ca(2+)](i) transients in CHO-M1 cells expressing heterologous rat M1 muscarinic subtype. The activation curve established by microfluorimetry in CHO-M1 cells using increasing concentrations of Bj-PRO-7a reached the maximum response in the presence of 3 μM Bj-PRO-7a (EC(50) = 0.25 ± 0.07 μM). The variation observed by calcium imaging in these cells ranged from 52 to 1218 nM (EC(50) = 0.31 ± 0.12 μM). [Ca(2+)](i) responses in CHO-M1 cells were largely inhibited by pirenzepine, a specific M1 antagonist. Neural-differentiated P19 cells expressing endogenous M1 receptors were also responsive to Bj-PRO-7a application, whereas no such response was observed in undifferentiated P19 cells not expressing muscarinic receptors. As further support for its specific action on M1 receptors, the peptide did not activate M3 subtypes in transfected CHO cells. Our findings provide a novel M1 muscarinic receptor agonist that could be used for basic research and even for pharmacological applications.
authors.affiliation: Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Priscilla D
authors.initials: PD
authors.lastname: Negraes
authors.forename: Claudiana
authors.initials: C
authors.lastname: Lameu
authors.forename: Mirian A F
authors.initials: MA
authors.lastname: Hayashi
authors.forename: Robson L
authors.initials: RL
authors.lastname: Melo
authors.forename: Antônio C M
authors.initials: AC
authors.lastname: Camargo
authors.forename: Henning
authors.initials: H
authors.lastname: Ulrich
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017837:Bothrops; D016466:CHO Cells; D020013:Calcium Signaling; D006224:Cricetinae; D003412:Cricetulus; D003435:Crotalid Venoms / Q000737:chemistry*; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology*; D051381:Rats; D043643:Receptor, Muscarinic M1 / Q000819:agonists* / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000819:agonists / Q000096:biosynthesis
chemical_list: D003435:Crotalid Venoms; D009842:Oligopeptides; D043643:Receptor, Muscarinic M1; D011994:Recombinant Proteins