STRUCTURAL AND FUNCTIONAL-ANALYSIS OF THE B-CELL EPITOPES RECOGNIZED BY ANTIRECEPTOR AUTOANTIBODIES IN PATIENTS WITH CHAGAS-DISEASE

IgG fractions of patients were screened for autoantibodies against the beta(1)- and beta(2)-adrenoceptors and the M2 acetylcholine receptor by enzyme immunoassays and surface plasmon resonance (SPR) using pepti des corresponding to the second extracellular loop of these receptors, A high prevalence of anti-M2 acetylcholine receptor and, in decreasin g order, of anti-beta(1)- and anti-beta(2)-adrenoceptor autoantibodies was shown. The enzyme immunoassays and the SPR studies on the anti-be ta(1) adrenoceptor and the M2 acetylcholine receptor autoantibodies we re dependent on the ionic strength of the interaction buffer, suggesti ng the importance of electrostatic interactions in Ab recognition. IgG fractions showed chronotropic effects on neonatal rat cardiomyocytes in vitro. The positive chronotropic effect was enhanced in the presenc e of 1 mu M of atropine, demonstrating a muscarinic stimulation by the IgG fractions in the presence of a beta-adrenergic stimulation, which was blocked by the use of 1 mu M of the beta(1)-selective antagonist bisoprolol. The beta(2)-selective antagonist ICI 118,551 only partiall y inhibited the positive chronotropic effect induced by the IgG fracti ons, confirming the minor functional importance of autoantibodies agai nst the beta(2)-adrenoceptor. Affinity-purified Abs confirmed that Abs against the beta(1)-adrenoceptors and the M2 muscarinic receptors exi st together with an Ab population recognizing a cross-reactive epitope on both receptors. This epitope could be identified as a polyanionic stretch present in the second extracellular loop of both the beta(1)-a drenoceptor and the M2 acetylcholine receptor, This stretch correspond s to the previously determined cross-reactive epitope between the PO r ibosomal protein of Trypanosoma cruzi and the beta(1)-adrenoceptor.