Photoaffinity Labeling of the Guinea Pig Pulmonary Mast Cell Beta-adrenergic Receptor

Abstract

Beta-adrenergic agonists can prevent mediator release from guinea pig pulmonary mast cells. By pharmacologic characterization, this response is mediated through a beta-2 receptor. Structural characterization of this receptor on the lung mast cell, however, has been limited by methods for isolation of this pulmonary cell. In this study, the guinea pig lung mast cell was isolated to greater than 90% purity, and its beta-adrenergic receptor identified by photoaffinity labeling with [125I]iodoazidobenzylpindolol (125IABP) and separation of membrane proteins by SDS-PAGE. We found the guinea pig pulmonary mast cell beta-adrenergic receptor to electrophorese as a heterogeneous protein between 68 and 116 kD. Photoaffinity labeling with 125IABP was protectable by alprenolol and isoproterenol but not by phentolamine and norepinephrine. Using subtype-selective compounds, the pulmonary mast cell receptor was established to be of a beta-2 subtype. This is the first report of the structural identification of a lung mast cell beta-adrenergic receptor and the first report of a beta-adrenergic receptor of approximately 100 kD in mass. This mast cell receptor is considerably larger than the 65 kD beta-adrenergic receptors that have been identified in whole lung and other tissues. Data we have obtained using Northern blot analysis of mast cell RNA suggest a protein message of 45 kD for this beta-adrenergic receptor and a high degree of glycosylation most likely accounts for the large molecular size observed.