title: Direct activation of gastric H,K-ATPase by N-terminal protein kinase C phosphorylation. Comparison of the acute regulation mechanisms of H,K-ATPase and Na,K-ATPase.
abstract: In this study we compared the protein kinase dependent regulation of gastric H,K-ATPase and Na,K-ATPase. The protein kinase A/protein kinase C (PKA/PKC) phosphorylation profile of H,K-ATPase was very similar to the one found in the Na,K-ATPase. PKC phosphorylation was taking place in the N-terminal part of the alpha-subunit with a stoichiometry of approximately 0.6 mol Pi/mole alpha-subunit. PKA phosphorylation was in the C-terminal part and required detergent, as is also found for the Na,K-ATPase. The stoichiometry of PKA-induced phosphorylation was approximately 0.7 mol Pi/mole alpha-subunit. Controlled proteolysis of the N-terminus abolished PKC phosphorylation of native H,K-ATPase. However, after detergent treatment additional C-terminal PKC sites became exposed located at the beginning of the M5M6 hairpin and at the cytoplasmic L89 loop close to the inner face of the plasma membrane. N-terminal PKC phosphorylation of native H,K-ATPase alpha-subunit was found to stimulate the maximal enzyme activity by 40-80% at saturating ATP, depending on pH. Thus, a direct modulation of enzyme activity by PKC phosphorylation could be demonstrated that may be additional to the well-known regulation of acid secretion by recruitment of H,K-ATPase to the apical membranes of the parietal cells. Moreover, a distinct difference in the regulation of H,K-ATPase and Na,K-ATPase is the apparent absence of any small regulatory proteins associated with the H,K-ATPase.
authors.affiliation: Department of Biophysics, University of Aarhus, Denmark. fc@biophys.au.dk
authors.forename: Flemming
authors.initials: F
authors.lastname: Cornelius
authors.forename: Yasser A
authors.initials: YA
authors.lastname: Mahmmoud
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis; D002478:Cells, Cultured; D004789:Enzyme Activation; D005753:Gastric Mucosa / Q000201:enzymology* / Q000378:metabolism; D017506:H(+)-K(+)-Exchanging ATPase / Q000737:chemistry / Q000378:metabolism*; D006706:Homeostasis / Q000502:physiology; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D010295:Parietal Cells, Gastric / Q000201:enzymology / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000737:chemistry / Q000378:metabolism*; D021122:Protein Subunits; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry / Q000378:metabolism*; D013552:Swine
chemical_list: D009097:Multienzyme Complexes; D021122:Protein Subunits; D011493:Protein Kinase C; D017506:H(+)-K(+)-Exchanging ATPase; D000254:Sodium-Potassium-Exchanging ATPase