Species-specific pharmacology of maximakinin, an amphibian homologue of bradykinin: putative prodrug activity at the human B2 receptor and peptidase resistance in rat
- Published
- Accepted
- Subject Areas
- Cardiology, Pharmacology
- Keywords
- bradykinin B2 receptor, maximakinin, MRGPRX2, human umbilical vein, angiotensin converting enzyme, in vivo hemodynamics
- Copyright
- © 2016 Charest-Morin et al.
- Licence
- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
- Cite this article
- 2016. Species-specific pharmacology of maximakinin, an amphibian homologue of bradykinin: putative prodrug activity at the human B2 receptor and peptidase resistance in rat. PeerJ Preprints 4:e2641v1 https://doi.org/10.7287/peerj.preprints.2641v1
Abstract
Maximakinin (MK), an amphibian peptide possessing the C-terminal sequence of bradykinin (BK), is a BK B2 receptor (B2R) agonist eliciting prolonged signaling. We reinvestigated this 19-mer for species-specific pharmacologic profile, in vivo confirmation of resistance to inactivation by angiotensin converting enzyme (ACE), value as a module for the design of fusion proteins that bind to the B2R in mammalian species and potential activity as a histamine releaser. Competition of the binding of [3H]BK to recombinant human myc-B2Rs in cells that express these receptors revealed that MK possessed a tenuous fraction (<0.1%) of the affinity of BK, despite being only ~10-fold less potent than BK in a contractility assay based on the human isolated umbilical vein. These findings are reconciled by the generation of C-terminal fragments, like Lys-Gly-Pro-BK and Gly-Pro-BK, when the latent MK is incubated with human venous tissue (LC-MS), supporting activation via hydrolysis upstream of the BK sequence. At the rat recombinant myc-B2R, MK had a lesser affinity than that of BK, but with a narrower margin (6.2-fold, radioligand binding competition). Accordingly, MK (10 nM) stimulated calcium transients in cells that expressed the rat receptors, but not the human B2R. Recombinant MRGPRX2, a receptor that mediates cationic peptide-induced mast cell secretion, minimally responded by increased [Ca+2]i to MK at 10 μM. Enhanced green fluorescent protein fused to MK (EGFP-MK) labeled cells that expressed rat, but not human B2Rs. Intravenous MK induced dose-dependent hypotensive, vasodilator and tachycardic responses in anesthetized rats and the effects were antagonized by pretreatment with icatibant but not modified by pyrilamine or enalaprilat. Strong species-specific responses to the toxin-derived peptide MK and its prodrug status in the isolated human vein were evidenced. Accordingly, MK in the EGFP-MK fusion protein is a pharmacophore module that confers affinity for the rat B2R, but not for the human form of the B2R. MK is unlikely to be an efficient mast cell activator, but its resistance to inactivation by ACE was confirmed in vivo.
Author Comment
This is a submission to PeerJ for review.
Supplemental Information
Confirmation of the sequence of 2 C-terminal fragments of MK identified in samples of MK incubated for 15 min in the presence of a human umbilical vein ring via tandem mass spectroscopy (LC-MS/MS)
The found N-terminal fragments of “n” residues (bn, in red) or of C-terminal ones (yn, in blue) of the postulated MK fragments are indicated along the mass/charge scale, thus confirming the identity of the parent peptides Gly-Pro-BK (A) and Lys-Gly-Pro-BK (B).
Raw data
Individual replicates for each averaged data point in figs. 1-3, 7 and 8.