N-terminal extended conjugates of the agonists and antagonists of both bradykinin receptor subtypes: Structure–activity relationship, cell imaging using ligands conjugated with fluorophores and prospect for functionally active cargoes

Abstract

N-terminal extended conjugates of the agonists and antagonists of both bradykinin receptor subtypes: structure-activity relationship, cell imaging using ligands conjugated with fluorophores and prospect for functionally active cargoes (3as,7as)-octahydroindole-2-carboxylic acid; Tic: 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 3  

Inhibitory effects of cytoskeleton disrupting drugs and GDP-locked Rab mutants on bradykinin B2 receptor cycling

“…The third element of the internalized complex organized around the B 2 R is the agonist ligand. FTC-B-9972, a recently described fluorescent B 2 R agonist resistant to endosomal breakdown [4], was exploited in HEK 293a cells that transiently expressed myc-B 2 R (Fig. 4).…”

“…Thus, the B 2 R combines with either type of non-visual -arrestins and the ligand-B 2 R--arrestin complex gets translocated into endosomes [1,2]. The fusion protein B 2 Rgreen fluorescent protein (B 2 R-GFP) -arrestins conjugated to fluorescent proteins and BK sequences that were extended with fluorophores were instrumental in modeling the BK-induced endocytosis, but also its recycling to the cell surface and the persistence of the ternary agonistreceptor--arrestin complexes in intracellular structures [3][4][5] (and literature cited herein).…”

“…Fluorescent BK analogs are rapidly transported by the B 2 R into Rab5-and early endosome autoantigen 1 (EEA1)-positive early endosomes and are later sorted in Rab7-positive late endosomes [3,4]. However, the BK-stimulated B 2 Rs are essentially completely recycled to the cell surface as a function of time, unless the agonist has an altered structure that confers resistance to proteolytic breakdown.…”

Vasopeptidase-activated latent ligands of the histamine receptor-1

“…However, we reported lately that the peptide Met-Lys-bradykinin-Ser-Ser is paradoxically activated by its reaction with ACE, that frees the BK C-terminal sequence needed to activate the cognate B 2 receptor (B 2 R) [2]. The modulatory role of vascular aminopeptidase N (APN) has also been illustrated, as the blockade of this ectopeptidase, expressed in vascular smooth muscle, potentiates such peptides as Lys-des-Arg 9 -BK (the optimal agonist of human and rabbit BK B 1 receptor), some peptide antagonists of B 1 receptors and angiotensin III [3,4].…”

“…Endosomal BK degradation obligatorily precedes the dissociation of β-arrestins from the B 2 R and receptor recycling to the cell surface, as shown by several inactivation-resistant B 2 R agonists that promote the persistence of this intracellular complex for at least 12 h and prolonged signaling [6,7]. Fluorophore conjugated analogs (carboxyfluorescein-or AlexaFluor-350-ε-aminocaproyl-BK) model the intracellular inactivation of the kinin, notably because the inhibitor of the proton pump V-ATPase, bafilomycin A1, prevents the time-dependent disappearance of the fluorescent peptides in endosomes of B 2 Rexpressing cells [8,9]. Free carboxyfluorescein is also released into the cytosol as a function of time in these cells, suggesting a particular strategy to release a drug cargo from BK conjugates.…”