From agonist to antagonist: Fab fragments of an agonist-like monoclonal anti-beta2-adrenoceptor antibody behave as antagonists

Item Type:	Article
Title:	From agonist to antagonist: Fab fragments of an agonist-like monoclonal anti-beta2-adrenoceptor antibody behave as antagonists
Creators Name:	Mijares, A. and Lebesgue, D. and Wallukat, G. and Hoebeke, J.
Abstract:	We previously demonstrated that the monoclonal antibody Mab6H8 raised against the second extracellular loop of the {beta}2-adrenoceptor ({beta}2-AR) had an agonist-like activity, mediated by the activation of L-type Ca2+ channels by protein kinase A through the adenylyl cyclase pathway. We suggested that this Mab acts by stabilizing an active dimeric conformation of the {beta}2-AR. To substantiate this hypothesis, we prepared monomeric Fab fragments of Mab6H8. Comparison of the physico-chemical parameters of antigen interaction with both the Mab and its Fab fragments were determined by surface plasmon resonance, showing a 5- to 10-fold lower affinity of the fragments compared with the bivalent antibody. We determined the biological activity of antibody and Fab fragments in two systems: spontaneous beating neonatal rat cardiomyocytes to study the chronotropic effects and isolated guinea pig cardiomyocytes to study L-type Ca2+ channel activation. Fab fragments as such had no 'agonist-like' effects in both systems but inhibited receptor activation with the {beta}2-specific agonist clenbuterol. Addition of a cross-linking rabbit anti-mouse IgG restored the agonist-like effect of the Fab fragments. These results suggest that Fab fragments induce a conformational change in the receptor, inhibiting the accessibility of the pharmacophore pocket to clenbuterol. Dimerization of this receptor conformation induces an agonist-like effect. Antireceptor antibodies can thus act both as agonist in the dimeric state and as antagonist in the monomeric state.
Keywords:	Adrenergic Beta-Antagonists, Beta-2 Adrenergic Receptors, Calcium, Cytosol, Heart, Immunoglobulin Fab Fragments, L-Type Calcium Channels, Monoclonal Antibodies, Myocardium, Wistar Rats, Animals, Guinea Pigs, Rats
Source:	Molecular Pharmacology
ISSN:	0026-895X
Publisher:	Amer Soc Pharmacology Experiamental Therapeutics
Volume:	58
Number:	2
Page Range:	373-379
Date:	1 August 2000
Official Publication:	http://molpharm.aspetjournals.org/cgi/content/abstract/58/2/373
PubMed:	View item in PubMed

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