Biochemical Studies of Structure, Function and Regulation of Gastric (H^+ + K^+)-ATPase

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The human gastric (H^+ + K^+)-ATPase gene (15 kilobases) was cloned, and its nucleotide sequence was determined. The gene has 22 exons and codes a protein of 1,035 residues including the initiator methionine (Mr=114,047). The phosphorylation site and pyridoxal 5'-phosphate- and fluorescein isothiocyanate-binding residues found in the rat and pig enzymes are also conserved in the human enzymes. The positions of introns in the human (H^+ + K^+)-ATPase gene are essentially the same as those in the human (Na^+ + K^+)-ATPase alpha and alpha III subunits. The similarity in organization of these two ATPase genes and the homology in the primary structures of their proteins (about 60%) suggest that these two genes were derived from a common ancestral gene. However, the 5'-flanking regions of the genes for (H^+ + K^+)-ATPase and the (Na^+ + K^+)-ATPase alpha (+) subunit show no apparent sequence homology, indicating that their transcriptions are requlated differently. The control region of the fast-twitch sarcoplasmic reticulum Ca^<2+>-ATPase gene also showed no sequence homology to that of (H^+ + K^+)-ATPase. The 5'-flanking region of the (H^+ + K^+)-ATPase gene contains potential binding sites for RNA polymerase II and various transcriptional regulation factors and several direct and inverted repeat sequences which may be important for specific and controlled expression of the genesin gastric parietal cells. We also cloned the rat gastric (H^+ + K^+)-ATPase gene. Comparison of its 5'-upstream region with the corresponding part of the human gene indicated the presence of highly conserved sequences. Nuclear proteins from gastric mucosa but not from liver recognizes these sequences. Such proteins must positively regulate gastric specific transcription, since Northern blot hybridization indicated that the (H^+ + K^+)-ATPase gene was transcribed only in gastric mucosa.