ALPHA-SUBUNIT AND BETA-SUBUNIT OF THE GASTRIC H+ K+-ATPASE ARE CONCORDANTLY TARGETED BY PARIETAL-CELL AUTOANTIBODIES ASSOCIATED WITH AUTOIMMUNE GASTRITIS/

We have previously shown that parietal cell autoantibodies predominant ly react with a 60-90 kDa gastric autoantigen(1,2), subsequently ident ified as the beta subunit of the gastric H+/K+-ATPase (EC 3.6.1.3) (pr oton pump)(3-5) whereas Karlsson et al(6) showed that these autoantibo dies primarily target the 95 kDa a subunit of the pump. In view of the se discordant results, we have reassessed the reactivity of parietal c ell autoantibodies with the two subunits of the gastric H+/K+-ATPase. We show here that all 26 parietal cell autoantibody-positive sera immu noblot both subunits under appropriate, but mutually exclusive, condit ions. Thus, reactivity of anti- parietal cell autoantibodies with the 95 kDa alpha subunit is optimal when the SDS-PAGE is carried out with samples which are reduced but not boiled. Whereas reactivity with the 60-90 kDa beta subunit is optimal with samples which are boiled but no t reduced. Autoantibody reactivity with the beta subunit is critically dependent on the presence of a full complement of N-linked glycans si nce partially deglycosylated protein, and recombinant beta subunit exp ressed in COS cells, bearing high mannose N-glycans, failed to bind to the autoantibody. These studies also suggest that B cell auto-epitope s are located on the lumenal domain of the beta subunit. Reactivity of parietal cell autoantibodies with a bacterial fusion protein incorpor ating the catalytic cytoplasmic domain of the a subunit suggests the p resence of auto-epitopes in this region of the molecule.