THE GASTRIC ACID PUMP AS A TARGET FOR ULCER TREATMENT

Summary

Principal Investigator: G Sachs
Affiliation: University of California
Country: USA
Abstract: The gastric H+, K+ ATPase catalyzes the final step of gastric acid secretion thereby generating a proton gradient across the canalicular membrane of greater than a million fold. It has been a focus of this laboratory for 30 years. Our interest in recent years has been the structure-function relationships of this pump and the mechanism of inhibition of this pump by covalent inhibitors (the proton pump inhibitors, PPIs) and the K+ -competitive reversible inhibitors (the acid pump antagonists, APAs).In the proposed studies, we plan to continue our site-directed mutagenesis approach coupled with detailed homology analysis and modeling using the 4 available crystal structures of the SERCA Ca ATPase as a template to better define transport by the gastric acid pump. The Ca ATPase although only 29% homologous to the H+-K+-ATPase, has a very similar overall structure and also uses carboxylic acid clusters in the membrane domain as the ion transport- binding and export-import sites as does the Na, K+ ATPase that is 75% homologous to the H+,K+ ATPase. We plan to delineate pathways for transport of hydronium ion from cytoplasm to lumen and K+ from lumen to cytoplasm by analyzing various enzyme activities including phosphorylation and dephosphorylation of selected site mutants. We have developed a hypothesis that the unique Lys791 insertion into one cluster of acids (D814, E820, E795) allows release of proton at the required pH~1.0 and K+ binding to luminal carbonyl groups displaces two of the carboxylic acids bound to lys 791, thereby allowing return ofIys791 and replacement by K+ at this site. New mutants will further define the ion transport pathways of the H+,K+ ATPase by homology modeling. Since acid- pump antagonists are in final clinical trials, we plan to define their site of binding to the enzyme more precisely by synthesizing a new class of compounds and identifying the amino acid residues whose mutation alters the affinity or nature of inhibition by these new compounds as we have done for the now classical imidazo-1,2a prydine class that often show unexpected negative side effects. Since an important step of acid secretory regulation involves a morphological transformation of the parietal cell wherein the ATPase moves from a cytoplasmic membrane location to the microvilli of the secretory canaliculus, we will continue our study of trafficking and sorting of the stably expressed subunit of the enzyme in polarized gastric cells. In addition, we will study the distribution of a YFP- subunit knock in- construct in the mouse stomach, living mouse gastric glands and in other tissuess such as the kidney where the enzyme is expressed but function is unknown. The scaffold proteins interacting with the (3 subunit will be elucidated using the the split ubiquitin method which is capable of defining those proteins associated with a particular membrane inserted protein. These studies will aid in clarifying the role of translocation of the pump in regulation of acid secretion. The results of the proposed research will further improve the agents used for the treatment of acid-related diseases and also our knowledge of the ATPase and cellular events involved in regulation of its activity.
Funding Period: 2000-09-15 - 2010-08-31
more information: NIH RePORT

Top Publications

  1. ncbi Use of the H,K-ATPase beta subunit to identify multiple sorting pathways for plasma membrane delivery in polarized cells
    Olga Vagin
    Department of Physiology, School of Medicine, UCLA and Veterans Administration Greater Los Angeles Health Care System, Los Angeles, California 90073, USA
    J Biol Chem 280:14741-54. 2005
  2. ncbi The gastric H,K ATPase as a drug target: past, present, and future
    George Sachs
    Department of Physiology, David Geffen School of Medicine, University of California at Los Angeles, CA, USA
    J Clin Gastroenterol 41:S226-42. 2007
  3. ncbi Identification of genomic targets downstream of p38 mitogen-activated protein kinase pathway mediating tumor necrosis factor-alpha signaling
    Cindy Zer
    Department of Physiology, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, California, USA
    Physiol Genomics 31:343-51. 2007
  4. ncbi The roles of the Na,K-ATPase beta 1 subunit in pump sorting and epithelial integrity
    Olga Vagin
    Department of Physiology, School of Medicine, UCLA, Los Angeles, CA 90073, USA
    J Bioenerg Biomembr 39:367-72. 2007
  5. ncbi Inverse correlation between the extent of N-glycan branching and intercellular adhesion in epithelia. Contribution of the Na,K-ATPase beta1 subunit
    Olga Vagin
    Department of Physiology, School of Medicine, University of California, Los Angeles, Veterans Administration Greater Los Angeles Health Care System, Los Angeles, California 90073, USA
    J Biol Chem 283:2192-202. 2008
  6. ncbi An ion gating mechanism of gastric H,K-ATPase based on molecular dynamics simulations
    Richard J Law
    Lawrence Livermore National Laboratory, Livermore, California 94550, USA
    Biophys J 95:2739-49. 2008
  7. ncbi Simple diagnostic tests to detect toxic alcohol intoxications
    Jai Moo Shin
    Medical and Research Services VHAGLA Healthcare System, UCLA Membrane Biology Laboratory, Division of Nephrology VHAGLA Healthcare System and David Geffen School of Medicine, Los Angeles, CA, USA
    Transl Res 152:194-201. 2008
  8. ncbi Role of N-glycosylation in trafficking of apical membrane proteins in epithelia
    Olga Vagin
    Department of Physiology, David Geffen School of Medicine at University of California, Bldg 113, Rm 324, 11301 Wilshire Blvd, Los Angeles, California 90073, USA
    Am J Physiol Renal Physiol 296:F459-69. 2009
  9. ncbi Pharmacology of proton pump inhibitors
    Jai Moo Shin
    Membrane Biology, David Geffen School of Medicine, University of California at Los Angeles, Room 324, Building 113, 11301 Wilshire Boulevard, Los Angeles, CA 90073, USA
    Curr Gastroenterol Rep 10:528-34. 2008
  10. ncbi Assembly with the Na,K-ATPase alpha(1) subunit is required for export of beta(1) and beta(2) subunits from the endoplasmic reticulum
    Elmira Tokhtaeva
    Department of Physiology, School of Medicine, UCLA and Veterans Administration Greater Los Angeles Health Care System, Los Angeles, USA
    Biochemistry 48:11421-31. 2009

Scientific Experts

Detail Information

Publications24

  1. ncbi Use of the H,K-ATPase beta subunit to identify multiple sorting pathways for plasma membrane delivery in polarized cells
    Olga Vagin
    Department of Physiology, School of Medicine, UCLA and Veterans Administration Greater Los Angeles Health Care System, Los Angeles, California 90073, USA
    J Biol Chem 280:14741-54. 2005
    ....
  2. ncbi The gastric H,K ATPase as a drug target: past, present, and future
    George Sachs
    Department of Physiology, David Geffen School of Medicine, University of California at Los Angeles, CA, USA
    J Clin Gastroenterol 41:S226-42. 2007
    ..The review concludes with a discussion of the mechanism of action and binding regions of a possible new class of drug for acid control, the K competitive acid pump antagonists...
  3. ncbi Identification of genomic targets downstream of p38 mitogen-activated protein kinase pathway mediating tumor necrosis factor-alpha signaling
    Cindy Zer
    Department of Physiology, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, California, USA
    Physiol Genomics 31:343-51. 2007
    ..Approximately one-third of the TNF-alpha-induced genes in FLS are regulated by the p38 MAPK signal pathway, showing that p38 MAPK is a possible target for suppressing proinflammatory gene expressions in rheumatoid arthritis...
  4. ncbi The roles of the Na,K-ATPase beta 1 subunit in pump sorting and epithelial integrity
    Olga Vagin
    Department of Physiology, School of Medicine, UCLA, Los Angeles, CA 90073, USA
    J Bioenerg Biomembr 39:367-72. 2007
    ..In addition, N-glycans contribute to the formation of cell-cell contacts between surface-attached dispersed cells by mediating lamellipodia formation and stabilizing the newly formed adherens junctions...
  5. ncbi Inverse correlation between the extent of N-glycan branching and intercellular adhesion in epithelia. Contribution of the Na,K-ATPase beta1 subunit
    Olga Vagin
    Department of Physiology, School of Medicine, University of California, Los Angeles, Veterans Administration Greater Los Angeles Health Care System, Los Angeles, California 90073, USA
    J Biol Chem 283:2192-202. 2008
    ..These results suggest epithelial cells can regulate tightness of cell junctions via remodeling of N-glycans, including those linked to the Na,K-ATPase beta(1)-subunit...
  6. ncbi An ion gating mechanism of gastric H,K-ATPase based on molecular dynamics simulations
    Richard J Law
    Lawrence Livermore National Laboratory, Livermore, California 94550, USA
    Biophys J 95:2739-49. 2008
    ..The movement of the M1M2 transmembrane segments, and the displacement of residues Q159, E160, Q110, and T152 during the conformational change, as well as the motions of E343 and L346, acted as the cytoplasmic-side gate...
  7. ncbi Simple diagnostic tests to detect toxic alcohol intoxications
    Jai Moo Shin
    Medical and Research Services VHAGLA Healthcare System, UCLA Membrane Biology Laboratory, Division of Nephrology VHAGLA Healthcare System and David Geffen School of Medicine, Los Angeles, CA, USA
    Transl Res 152:194-201. 2008
    ..The relatively high sensitivity and specificity of the tests as a whole will facilitate the rapid diagnosis of each of these alcohol intoxications...
  8. ncbi Role of N-glycosylation in trafficking of apical membrane proteins in epithelia
    Olga Vagin
    Department of Physiology, David Geffen School of Medicine at University of California, Bldg 113, Rm 324, 11301 Wilshire Blvd, Los Angeles, California 90073, USA
    Am J Physiol Renal Physiol 296:F459-69. 2009
    ..Finally, we review existing hypotheses on the mechanism of apical sorting and discuss the potential roles of the lectins, VIP36 and galectin-3, as putative apical sorting receptors...
  9. ncbi Pharmacology of proton pump inhibitors
    Jai Moo Shin
    Membrane Biology, David Geffen School of Medicine, University of California at Los Angeles, Room 324, Building 113, 11301 Wilshire Boulevard, Los Angeles, CA 90073, USA
    Curr Gastroenterol Rep 10:528-34. 2008
    ..PPIs with longer half-life promise to improve acid suppression. All PPIs give excellent healing of peptic ulcers and produce good results in reflux esophagitis. PPIs combined with antibiotics eradicate Helicobacter pylori...
  10. ncbi Assembly with the Na,K-ATPase alpha(1) subunit is required for export of beta(1) and beta(2) subunits from the endoplasmic reticulum
    Elmira Tokhtaeva
    Department of Physiology, School of Medicine, UCLA and Veterans Administration Greater Los Angeles Health Care System, Los Angeles, USA
    Biochemistry 48:11421-31. 2009
    ....
  11. ncbi Cytoplasmic histidine kinase (HP0244)-regulated assembly of urease with UreI, a channel for urea and its metabolites, CO2, NH3, and NH4(+), is necessary for acid survival of Helicobacter pylori
    David R Scott
    Department of Physiology, David Geffen School of Medicine at UCLA, 405 Hilgard Ave, Los Angeles, California 90024, USA
    J Bacteriol 192:94-103. 2010
    ..Assembly of a pH-regulatory complex of active urease with UreI provides an advantage for periplasmic buffering...
  12. ncbi Analysis of the gastric H,K ATPase for ion pathways and inhibitor binding sites
    Keith Munson
    Laboratory of Membrane Biology, David Geffen School of Medicine at UCLA, and VA GLAHS, Los Angeles, California 90073, USA
    Biochemistry 46:5398-417. 2007
    ..Finally, the expanded luminal vestibule of the E2P model explains high-affinity ouabain binding in a mutant of the H,K ATPase [Qiu et al. (2005) J. Biol. Chem. 280, 32349-32355]...
  13. ncbi Urea transport in bacteria: acid acclimation by gastric Helicobacter spp
    G Sachs
    Department of Physiology, Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90073, USA
    J Membr Biol 212:71-82. 2006
    ..The finding also of upregulation of the two-component histidine kinase HP0165 and its response element HP0166, illustrates the complexity of the acid acclimation processes involved in gastric colonization by this pathogen...
  14. ncbi Inhibitor and ion binding sites on the gastric H,K-ATPase
    Keith Munson
    Geffen School of Medicine, University of California at Los Angeles, and VAGLAHS, Los Angeles, California 90073, USA. kmunson@ ucla.edu
    Biochemistry 44:5267-84. 2005
    ..This site of K(+) binding is predicted to destabilize hydrogen bonds between these carboxylates and the -NH(3)(+) group of Lys791, allowing the Lys791 side chain to return to its E(1) position...
  15. ncbi Topography of the membrane domain of the liver Na+-dependent bile acid transporter
    Olga Mareninova
    Department of Physiology, David Geffen School of Medicine, University of California at Los Angeles, and VA Greater Los Angeles Healthcare System, Los Angeles, California 90073, USA
    Biochemistry 44:13702-12. 2005
    ..This new method provided evidence for seven membrane segments giving a new model of the membrane domain of this protein and probably the homologous ileal transporter, with H7/H8 as the transport region...
  16. ncbi Recombinant addition of N-glycosylation sites to the basolateral Na,K-ATPase beta1 subunit results in its clustering in caveolae and apical sorting in HGT-1 cells
    Olga Vagin
    Department of Physiology, School of Medicine at UCLA, Los Angeles, California 90073, USA
    J Biol Chem 280:43159-67. 2005
    ....
  17. ncbi Acid acclimation by Helicobacter pylori
    George Sachs
    Laboratory of Membrane Biology, David Geffen School of Medicine at the University of California Los Angeles, USA
    Physiology (Bethesda) 20:429-38. 2005
    ..1 in acidic media, whereas other neutralophiles cannot regulate periplasmic pH and thus only transit the stomach...
  18. ncbi Functional consequences of the oligomeric form of the membrane-bound gastric H,K-ATPase
    Jai Moo Shin
    Department of Physiology and Medicine, University of California at Los Angeles, and VA Greater Los Angeles Healthcare System, Los Angeles, California 90073, USA jaishin ucla edu
    Biochemistry 44:16321-32. 2005
    ..It appears that the catalytic subunits of the oligomer during turnover in intact gastric vesicles are restricted to a reciprocal E(1):E(2) configuration...
  19. ncbi Transcriptomes of purified gastric ECL and parietal cells: identification of a novel pathway regulating acid secretion
    Nils W G Lambrecht
    Department of Pathology, David Geffen School of Medicine, University of California Los Angeles, California, USA
    Physiol Genomics 25:153-65. 2006
    ....
  20. ncbi Characterization of the inhibitory activity of tenatoprazole on the gastric H+,K+ -ATPase in vitro and in vivo
    Jai Moo Shin
    Department of Physiology and Medicine, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA, USA
    Biochem Pharmacol 71:837-49. 2006
    ....
  21. ncbi Involvement of the HP0165-HP0166 two-component system in expression of some acidic-pH-upregulated genes of Helicobacter pylori
    Yi Wen
    Membrane Biology Laboratory, Department of Physiology, David Geffen School of Medicine at UCLA, VA Greater Los Angeles Healthcare System, Los Angeles, California 90073, USA
    J Bacteriol 188:1750-61. 2006
    ..Regulation of these genes responds either to a decrease in the periplasmic pH alone (HP0165 dependent) or also to a decrease in the cytoplasmic pH (HP0165 independent)...
  22. ncbi The role of the beta1 subunit of the Na,K-ATPase and its glycosylation in cell-cell adhesion
    Olga Vagin
    Department of Physiology, School of Medicine, UCLA and Veterans Affairs Greater Los Angeles Health Care System, Los Angeles, California 90073, USA
    J Biol Chem 281:39573-87. 2006
    ..Further, normal glycosylation of the Na,K-ATPase beta(1) subunit is essential for the stable association of the pump with the adherens junctions and plays an important role in cell-cell contact formation...
  23. ncbi The HP0165-HP0166 two-component system (ArsRS) regulates acid-induced expression of HP1186 alpha-carbonic anhydrase in Helicobacter pylori by activating the pH-dependent promoter
    Yi Wen
    The Membrane Biology Laboratory, Department of Physiology, David Geffen School of Medicine at UCLA, USA
    J Bacteriol 189:2426-34. 2007
    ....
  24. ncbi 1-Arylsulfonyl-2-(pyridylmethylsulfinyl) benzimidazoles as new proton pump inhibitor prodrugs
    Jai Moo Shin
    Department of Physiology, David Geffen School of Medicine, University of California at Los Angeles, and VA Greater Los Angeles Healthcare System, Los Angeles, CA 90073, USA
    Molecules 14:5247-80. 2009
    ..New arylsulfonyl proton pump inhibitor (PPI) prodrug forms were synthesized. These prodrugs provided longer residence time of an effective PPI plasma concentration, resulting in better gastric acid inhibition...