A Chimeric Gastric H+,K+-ATPase Inhibitable with Both Ouabain and SCH 28080

Asano, Shinji; Matsuda, Saiko; Hoshina, Satomi; Sakamoto, Shinya; Takeguchi, Noriaki

doi:10.1074/jbc.274.11.6848

Cited by 30 publications

(28 citation statements)

References 34 publications

(30 reference statements)

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“…The crystallographic data may, however, predict an arrangement of the ten α1-subunit helices within the plane of the lipid bilayer that is common to both enzymes. In a third approach, chimeras of individual Na/KATPase loop regions were inserted into either the skeletal muscle sarcoplasmic reticulum/ endoplasmic reticulum Ca 2+ -ATPase (SERCA1) [39,40] or the gastric H/K-ATPase [41,42]. The results suggest that M1-M2, M3-M4 and M5-M6 extracellular loops (see Figure 2) are important for ouabain sensitivity.…”

Section: The Ouabain Binding Site On the α1-subunitmentioning

confidence: 99%

Progesterone binding to the α1-subunit of the Na/K-ATPase on the cell surface: Insights from computational modeling

2008

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Progesterone triggers the resumption of meiosis in the amphibian oocyte through a signaling system at the plasma membrane. Analysis of [ 3 H]ouabain and [ 3 H]progesterone binding to the plasma membrane of the Rana pipiens oocyte indicates that progesterone competes with ouabain for a low affinity ouabain binding site on a 112 kDa α1-subunit of the membrane Na/K-ATPase. Published amino acid sequences from both low and high affinity ouabain binding α1-subunits are compared, together with published site-directed mutagenesis studies of ouabain binding. We propose that the progesterone binding site is located in the external loop (23 amino acids) between the M1-M2 transmembrane helices. Analysis of loop topology and the countercurrent hydrophobicity/polarity gradients within the M1-M2 loop further suggest that the polar β and hydrophobic α surfaces of the planar progesterone molecule interact with opposite sides of the amino acid loop. The 19-angular methyl group of progesterone is essential for activity; it could bind to the C-terminal region of the M1-M2 loop. Maximum biological activity requires formation of hydrogen-bond networks between the 3-keto group of progesterone and Arg 118 , Asp 129 and possibly Glu 122-124 in the C-terminal region of the loop. The 20-keto group hydrogen may in turn hydrogen bond to Cys 111 near the M1 helix. Peptide flexibility undergoes a maximal transition near the midway point in the M1-M2 loop, suggesting that folding occurs within the loop, which further stabilizes progesterone binding.

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Section: The Ouabain Binding Site On the α1-subunitmentioning

confidence: 99%

Progesterone binding to the α1-subunit of the Na/K-ATPase on the cell surface: Insights from computational modeling

2008

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“…The cardiac glycoside binding site is presumably located within the fiveextracellular loops (TM1-TM2, TM3-TM4, etc.). Studies utilizing chimeras of individual Na + , K + -ATPase loop regions inserted into either the skeletal muscle sarcoplasmic reticulum/endoplasmic reticulum Ca 2+ -ATPase (SERCA1a) [7,8] or the gastric H + , K + -ATPase [9,10] have identified the TM1-TM2, TM3-TM4 and TM5-TM6 extracellular loops as important for binding of the cardiac glycoside, ouabain. More precisely, mutagenesis studies have identified several amino acid residues (C104, Y108, Q111, P118, D121 and N122) found in the TM1 segment and TM1-TM2 loop as critically important for the high affinity binding of ouabain [11][12][13].…”

Section: Introductionmentioning

confidence: 99%

Elucidation of the Na+, K+-ATPase digitalis binding site

Keenan

DeLisle²,

Welsh

et al. 2005

Journal of Molecular Graphics and Modelling

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“…Although the first photoaffinity labeling study indicated that the first extracellular loop (the M1/M2 loop) is the direct binding site of SCH 28080 (19), our mutational and chimeric studies showed that the M1/M2 loop is not the direct binding site (6,20). Several mutants showed lower sensitivity to SCH 28080: the M336I and V339I mutants (mutants in Met 336 and Val 339 in the M4 segment), the K793S and E797D mutants (mutants in Lys 793 and Glu 797 in the M5 segment), and the C815T mutant (mutant in Cys 815 either in or close to the M6 segment) (21)(22)(23).…”

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confidence: 99%

The Cavity Structure for Docking the K+-competitive Inhibitors in the Gastric Proton Pump

Asano¹,

Yoshida

Yashiro

et al. 2004

Journal of Biological Chemistry

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2-Methyl-8-(phenylmethoxy)imidazo[1,2-a]pyridine-3-acetonitrile (SCH 28080) is a reversible inhibitor specific for the gastric proton pump. The inhibition pattern is competitive with K ؉ . Here we studied the binding sites of this inhibitor on the putative three-dimensional structure of the gastric proton pump ␣-subunit that was constructed by homology modeling based on the structure of sarcoplasmic reticulum Ca 2؉ pump. Alanine and serine mutants of Tyr 801 located in the fifth transmembrane segment of the gastric proton pump ␣-subunit retained the 86 Rb transport and K ؉ -dependent ATPase (K ؉ -ATPase) activities. These mutants showed 60 -80-times lower sensitivity to SCH 28080 than the wild type in the 86 Rb transport activity. The K ؉ -ATPase activities of these mutants were not completely inhibited by SCH 28080. The sensitivity to SCH 28080 was dependent on the bulkiness of the side chain at this position. Therefore, the side chain of Tyr 801 is important for the interaction with this inhibitor. In the three-dimensional structure of the E 2 form (conformation with high affinity for K ؉ ) of the gastric proton pump, Tyr 801 faces a cavity surrounded by the first, fourth, fifth, sixth, and eighth transmembrane segments and fifth/sixth, seventh/eighth, and ninth/tenth loops. SCH 28080 can dock in this cavity. However, SCH 28080 cannot dock in the same location in the E 1 form (conformation with high affinity for proton) of the gastric proton pump due to the drastic rearrangement of the transmembrane helices between the E 1 and E 2 forms. These results support the idea that this cavity is the binding pocket of SCH 28080.The gastric H ϩ ,K ϩ -ATPase is a proton pump that is responsible for gastric acid secretion (1). This pump consists of two kinds of subunits, the ␣-and ␤-subunits. The ␣-subunit is the catalytic subunit containing 10 transmembrane segments. The ATP binding site (2, 3) and the cation binding sites are located on this subunit (4 -10). The ␤-subunit is the non-catalytic glycoprotein containing a single transmembrane segment and is involved in stabilization as well as targeting of the ␣-subunit to the plasma membrane (11). This pump belongs to the family of type II P-type ATPases, which include sarcoplasmic reticulum (SR) 1 Ca 2ϩ -ATPase (Ca 2ϩ pump) and Na ϩ ,K ϩ -ATPase (Na ϩ pump) (12).At present, inhibitors specific for the gastric proton pump are classified into two groups. Irreversible inhibitors such as omeprazole, rabeprazole, and lansoprazole, termed the proton pump inhibitors, are the first group. They are activated in acidic lumens, modify the cysteine residues located on the luminal side of the proton pump, and inhibit its pump activity. They are clinically used for controlling hyperacidity. The second classified group of specific proton pump inhibitors are the reversible inhibitors such as SCH 28080 (13) and 3-amino-5-methyl-2-(2-methyl-3-thienyl)imidazo[1,2-a]thieno[3,2-c]pyridine (SPI-447) (Fig. 1) (14), which are described as acid pump antagonists (APAs). They bind to the E 2 or E 2...

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A Chimeric Gastric H+,K+-ATPase Inhibitable with Both Ouabain and SCH 28080

Cited by 30 publications

References 34 publications

Progesterone binding to the α1-subunit of the Na/K-ATPase on the cell surface: Insights from computational modeling

Progesterone binding to the α1-subunit of the Na/K-ATPase on the cell surface: Insights from computational modeling

Elucidation of the Na+, K+-ATPase digitalis binding site

The Cavity Structure for Docking the K+-competitive Inhibitors in the Gastric Proton Pump

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