title: Dimerization of the gastric H+, K(+)-ATPase.
abstract: When the gastric H+, K(+)-ATPase was solubilized by n-dodecyl beta-D-maltoside and electrophoresed in blue native-polyacrylamide gels (BN-PAGE), one major band at about 360 kDa was observed. Since this band was recognized by both monoclonal antibodies 1218 (anti-alpha) and wheat germ agglutinin (anti-beta), the H+, K(+)-ATPase in its native state exists in a dimeric (alpha beta)2 form. The site of interaction between the heterodimers was determined using Cu(2+)-phenanthroline cross-linking. The Cu(2+)-phenanthroline reagent reacted with the H+, K(2+)-ATPase activity. This cross-linking and enzyme inhibition were prevented by ATP. Cross-linking followed by N-ethylmaleimide blockade of maleimide-reactive SH groups, then reduction and fluorescein 5-maleimide labeling, then reduction and fluorescent tryptic peptide of about 6.5 kDa that had been cross-linked. Since its N-terminal amino acid is Val561, the peptide probably ends at Arg616 or Arg621 and Cys565 and/or Cys615 are probably within the region of closest contact between the two alpha-subunits.
authors.affiliation: Wadsworth Veterans Administration Medical Center, Los Angeles, California, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Shin
authors.forename: G
authors.initials: G
authors.lastname: Sachs
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003300:Copper; D003432:Cross-Linking Reagents / Q000737:chemistry; D003545:Cysteine / Q000737:chemistry; D004791:Enzyme Inhibitors; D005753:Gastric Mucosa / Q000201:enzymology*; D017506:H(+)-K(+)-Exchanging ATPase / Q000737:chemistry*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010618:Phenanthrolines / Q000737:chemistry / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D054328:Proton Pump Inhibitors; D013552:Swine
chemical_list: D003432:Cross-Linking Reagents; D004791:Enzyme Inhibitors; D010618:Phenanthrolines; D054328:Proton Pump Inhibitors; D003300:Copper; D017506:H(+)-K(+)-Exchanging ATPase; D003545:Cysteine