Chapter 2 Structure and function of gastric H,K-ATPase

Catalytic Function of Nongastric H,K-ATPase Expressed in Sf-21 Insect Cells

“…The Atp1al1−βHK complex was found to be sensitive to vanadate like all other known P-type ATPases ( − ); this inhibitor in 100 μM concentration completely abolished Atp1al1−βHK ATPase activity (data not shown). The Atp1al1−βHK ATPase activity was found to be resistant to oligomycin (5 μg/mL) and 1 mM NaN 3 or KN 3 (data not shown).…”

“…Previous studies of ion sensitivity of other X,K-ATPases have demonstrated that Na + also partially inhibits ATP hydrolysis by the gastric H,K-ATPase but at an alkaline pH and has little or no effect on the activity at pH ≤7.0, thus revealing the possibility of competition between protons and Na + on cytoplasmic sites ( , ). Sodium ions may also substitute for protons in the ion-transporting mechanism of the gastric H,K-ATPase at very low H + concentration (pH 8.5) but not at a neutral pH ( , ). In the case of Na,K-ATPase, K + has been shown to act as a competitive inhibitor of Na + binding at cytoplasmic sites, thus suggesting that Na + /K + antagonism could be a plausible mechanism of pump regulation (for review, see ref 53 ).…”

“…Potassium-dependent animal ATPases (X,K-ATPases) are the most closely related among the various P-type ATPases (). All of the known X,K-ATPases function as cation exchangers which pump K + into the cell and Na + or H + out of the cell, exhibiting a much higher level of sequence homology between their catalytic α-subunits (∼110 kDa) than with other P-ATPases and containing a glycosylated β-subunit (core protein ∼35 kDa) which is absent in other P-ATPases ( − ). The X,K-ATPase family combines three distinct groups of ion pumps.…”

“…The X,K-ATPase family combines three distinct groups of ion pumps. Two groups, one consisting of the Na,K-ATPase isozymes formed by four α and three β isoforms and the second which includes the gastric H,K-ATPase, have long been known and studied extensively ( , ). Recently discovered catalytic α-subunits of nongastric H,K-ATPases encoded by the human ATP1AL1 gene and its animal homologues represent the third distinct group ( − ).…”

Role of Negatively Charged Residues in the Fifth and Sixth Transmembrane Domains of the Catalytic Subunit of Gastric H+,K+-ATPase

“…This results in a more C-terminally located position of the transmembrane segments M5 and M6 in H ϩ ,K ϩ -ATPase as compared with the original models for Ca 2ϩ -ATPase and Na ϩ ,K ϩ -ATPase (16,17). Despite this uncertainty, a number of site-directed mutagenesis studies aimed at elucidating the function of these negatively charged residues in the M5-M6 region in Ca 2ϩ -ATPase of both the SERCA (3,5,(22)(23)(24) and the plasma membrane type (25) (18,21,30), only one study with mutants has been published until now (31), since functional expression of this enzyme system has only recently been successfully carried out (31)(32)(33). We report here mutational studies in which six negatively charged amino acid residues within or close to the fifth and sixth transmembrane segments of the catalytic subunit have been converted into the corresponding acid amides.…”