CHARACTERIZATION OF A BETA-SUBUNIT OF THE GASTRIC H+/K+-TRANSPORTING ATPASE

The catalytic subunit of the H+/K+-transporting ATPase (EC 3.6.1.3) has 62% identity to the α, or catalytic subunit, of the Na+/K+-transporting ATPase (EC 3.6.1.37); however, a homologous β subunit was unknown until recently. Removal of the carbohydrate from purified hog H+/K+ATPase vesicles reveals a 35-kDa peptide that, when fragmented with protease V8, gives sequences homologous to both β1 and β2 subunits of the Na+/K+-ATPase. cDNA clones for a β subunit of the gastric H+/K+-ATPase were isolated from a rabbit stomach cDNA library by using degenerate 17-mer oligonucleotide probes made to the protease V8-treated peptides. An open reading frame (54-926) encodes a predicted 291-amino acid peptide with M(r) = 33,320, which exhibits 31% and 44% homologies to the Na+/K+-ATPase β1 and Na+/K+-ATPase β2 proteins, respectively. A Kyte-Doolittle hydropathy plot predicts a single N-terminal transmembrane domain with a small hydrophobic region near the C terminus. The presumed extracytosolic domain contains seven potential N-linked glycosylation sites and six out of nine cysteines. Northern (RNA) blot analysis of stomach RNA with the rabbit H+/K+-ATPase β probe identifies a single mRNA of 1.3-1.5 kilobases, similar in concentration to the α subunit mRNA. The presence of a defined gastric H+/K+-ATPase β subunit extends the homology between H+/K+-ATPase and the Na+/K+-ATPase subclass of phosphoenzyme transport ATPases and distinguishes them from the monomeric Ca2+ and proton pump subclasses.