Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cDNA for ATP1AL1--the fifth member of the human Na,K-/H,K-ATPase gene family--was cloned and sequenced. The deduced primary ATP1AL1 translation product is 1,039 amino acids in length and has Mr of 114,543. The encoded protein has all of the structural features common to known catalytic subunits of P-type membrane ion-transporting ATPases and is equally distant (63-64% of identity) from the Na,K-ATPase isoforms and the gastric H,K-ATPase. The ATP1AL1 encoded protein was proposed to represent a new separate group within the family of human potassium-dependent ion pumps.
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PMID:Cloning and characterization of the entire cDNA encoded by ATP1AL1--a member of the human Na,K/H,K-ATPase gene family. 804 93

To study the role of the Na,K-ATPase beta subunit in the ion transport activity, we have coexpressed the Bufo alpha 1 subunit (alpha 1) with three different isotypes of beta subunits, the Bufo Na,K-ATPase beta 1 (beta 1NaK) or beta 3 (beta 3NaK) subunit or the beta subunit of the rabbit gastric H,K-ATPase (beta HK), by cRNA injection in Xenopus oocyte. We studied the K+ activation kinetics by measuring the Na,K-pump current induced by external K+ under voltage clamp conditions. The endogenous oocyte Na,K-ATPase was selectively inhibited, taking advantage of the large difference in ouabain sensitivity between Xenopus and Bufo Na,K pumps. The K+ half-activation constant (K1/2) was higher in the alpha 1 beta 3NaK than in the alpha 1 beta 1NaK groups in the presence of external Na+, but there was no significant difference in the absence of external Na+. Association of alpha 1 and beta HK subunits produced active Na,K pumps with a much lower apparent affinity for K+ both in the presence and in the absence of external Na+. The voltage dependence of the K1/2 for external K+ was similar with the three beta subunits. Our results indicate that the beta subunit has a significant influence on the ion transport activity of the Na,K pump. The small structural differences between the beta 1NaK and beta 3NaK subunits results in a difference of the apparent affinity for K+ that is measurable only in the presence of external Na+, and thus appears not to be directly related to the K+ binding site. In contrast, association of an alpha 1 subunit with a beta HK subunit results in a Na,K pump in which the K+ binding or translocating mechanisms are altered since the apparent affinity for external K+ is affected even in the absence of external Na+.
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PMID:Modulation of the Na,K-pump function by beta subunit isoforms. 805 80

The kidney plays an essential role in regulating potassium and acid balance. A major site for these regulations is in the collecting tubule. In the present study, we report the primary sequence of a novel alpha subunit of the P-ATPase gene family, which we isolated from the urinary bladder epithelium of the toad Bufo marinus, the amphibian equivalent of the mammalian collecting tubule. The cDNA encodes a protein of 1,042 amino acids which shares approximately 67% identity with the alpha 1 subunit of the ouabain-inhibitable Na,K-ATPase and approximately 69% identity with the alpha subunit of the SCH28080-inhibitable gastric H,K-ATPase. When coexpressed in Xenopus oocytes with a beta subunit isolated from the same cDNA library, the ATPase is able to transport rubidium (a potassium surrogate) inward, and hydrogen outward, leading to alkalization of the intracellular compartment and acidification of the external medium. The novel ATPase has a unique pharmacological profile showing intermediate sensitivity to both ouabain and SCH28080. Our findings indicate that the bladder ATPase is a member of a new ion motive P-ATPase subfamily. The bladder ATPase is expressed in the urinary tract but not in the stomach or the colon. This H,K-ATPase may be one of the molecules involved in H+ and K+ homeostasis, mediating the transport of these ions across urinary epithelia and therefore regulating their urinary excretion.
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PMID:Mechanisms of urinary K+ and H+ excretion: primary structure and functional expression of a novel H,K-ATPase. 825 41

The ubiquitous Na,K- and the gastric H,K-pumps are heterodimeric plasma membrane proteins composed of an alpha and a beta subunit. The H,K-ATPase beta subunit (beta HK) can partially act as a surrogate for the Na,K-ATPase beta subunit (beta NK) in the formation of functional Na,K-pumps (Horisberger et al., 1991. J. Biol. Chem. 257:10338-10343). We have examined the role of the transmembrane and/or the ectodomain of beta NK in (a) its ER retention in the absence of concomitant synthesis of Na,K-ATPase alpha subunits (alpha NK) and (b) the functional expression of Na,K-pumps at the cell surface and their activation by external K+. We have constructed chimeric proteins between Xenopus beta NK and rabbit beta HK by exchanging their NH2-terminal plus transmembrane domain with their COOH-terminal ectodomain (beta NK/HK, beta HK/NK). We have expressed these constructs with or without coexpression of alpha NK in the Xenopus oocyte. In the absence of alpha NK, Xenopus beta NK and all chimera that contained the ectodomain of beta NK were retained in the ER while beta HK and all chimera with the ectodomain of beta HK could leave the ER suggesting that ER retention of unassembled Xenopus beta NK is mediated by a retention signal in the ectodomain. When coexpressed with alpha NK, only beta NK and beta NK/HK chimera assembled efficiently with alpha NK leading to similar high expression of functional Na,K-pumps at the cell surface that exhibited, however, a different apparent K+ affinity. beta HK or chimera with the transmembrane domain of beta HK assembled less efficiently with alpha NK leading to lower expression of functional Na,K-pumps with a different apparent K+ affinity. The data indicate that the transmembrane domain of beta NK is important for efficient assembly with alpha NK and that both the transmembrane and the ectodomain of beta subunits play a role in modulating the transport activity of Na,K-pumps.
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PMID:Role of the transmembrane and extracytoplasmic domain of beta subunits in subunit assembly, intracellular transport, and functional expression of Na,K-pumps. 827 95

To determine if proton translocating adenosine triphosphatases (ATPases) can be localized at the blood-brain barrier, isolated rat cerebral microvessels and cerebral synaptosomal preparations were assayed for ATPase activity in the presence of various inhibitors. N-ethylmaleimide-sensitivity could be consistently found in both cerebral microvessel and synaptosomal preparations. There was no vanadate sensitive component in the presence of ouabain, oligomycin and EGTA. Immunoblotting of cerebral microvessels and synaptosomes with a monoclonal antibody (E11) against the 31 kDa subunit of the vacuolar type H-ATPase pump identified a discrete 31 kDa band. Diffuse immunocytochemical staining of cerebral cortical tissue, predominantly in choroid plexus, could be found with E11 but not with HK alpha N2, an H,K-ATPase specific antibody, nor with a non-specific mouse monoclonal antibody (MOPC-21). Immunoblotting with HK alpha N2 showed an immunoreactive 76 kDa band, not present with the preimmune serum or the antibody preabsorbed with the immunizing synthetic peptide. It is concluded that the vascular type H-ATPase and not the gastric H,K-ATPase is present in cerebral tissue including cerebral microvessels and choroid plexus. Non-specific immunoreactivity may account for the 76 kDa band observed in the immunoblots using the HK alpha N2 antibody although presence of a degradation product of H,K-ATPase can not be ruled out. The functional role of the vacuolar H-ATPase in the blood-brain barrier remains to be determined.
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PMID:Identification of proton-translocating adenosine triphosphatases in rat cerebral microvessels. 828 67

Tertiary amines like imidazole and triallylamine lower the apparent affinity of K+ in the ATP hydrolysis reaction of pig gastric H,K-ATPase in a pH and amine concentration dependent way. The mechanism and sidedness of this effect was studied by analyzing the partial reactions of the enzyme in both leaky and ion-tight vesicles. In leaky vesicles Tris and Hepes had nearly no effect on the apparent Km for K+ in the ATPase reaction, but imidazole (Ki = 13 mM) and triallylamine (Ki = 1.6 mM) markedly decreased the K+ affinity. The steady-state ATP-phosphorylation level in the absence of K+ was not or only slightly affected by these compounds. The reduction of the ATP-phosphorylation level by K+, however, again depended on both the type and concentration of tertiary amine used. A comparable K(+)-amine antagonism was observed in the dephosphorylation reaction. In tightly sealed vesicles, where no activation of K+ at the luminal side could occur, K+ reduced the affinity for ATP in the phosphorylation reaction. Triallylamine counteracted this effect. The K(+)-activated p-nitrophenylphosphatase activity in these ion-tight vesicles also showed a K(+)-triallylamine antagonism. Inhibition of H,K-ATPase activity in these vesicles by triallylamine was immediate (with nigericin present in order to allow intravesicular K+ activation), suggesting the transmembrane feature of this inhibition. These results indicate that tertiary amines decrease the affinity for K+ at both luminal and cytosolic binding sites by interaction at the cytosolic side of the membrane. This results in shifts in the equilibrium of both the E1.H<==>E1.K transition and in the dephosphorylation reaction, E2-P-->E2.K.
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PMID:Tertiary amines as antagonists of both the luminal and cytosolic K(+)-site of gastric H,K-ATPase. 829 18

Cultured pig kidney epithelial cells were transfected with a chimeric P-type ATPase catalytic subunit composed of the NH2-terminal half of the rat gastric H,K-ATPase and the COOH-terminal half of the rat Na,K-ATPase (alpha 1 isoform). Low concentrations of ouabain (< or = 0.2 mM) were used to inhibit completely the endogenous pig Na,K-ATPase and high concentrations (5 nM) to test the sensitivity of the chimeric rodent pump. In the presence of a low concentration of ouabain, a small but significant inhibition of residual Rb+(K+) influx by 5 mM ouabain was observed in only the transfected cells. Conditions were found in which a similar component of Rb+ influx was inhibited by the gastric H,K-ATPase inhibitor SCH28080, consistent with SCH28080 binding to the extracellular H1-H2 loop of this enzyme. These experiments demonstrate that this chimera behaves as a functional ion pump and indicate that the protein domains involved in cardiac glycoside binding are not confined to the amino-terminal half of the Na,K-ATPase.
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PMID:Functional properties of an H,K-ATPase/Na,K-ATPase chimera. 838 26

The ion-transporting H,K-ATPase and Na,K-ATPase enzymes are each composed of an alpha and a beta subunit. It is known that assembly of the alpha and beta subunits of the Na,K-ATPase is necessary for the cell-surface delivery of the active enzyme. We have examined the molecular domains involved in the assembly of the H,K-ATPase and Na,K-ATPase alpha and beta subunits by expressing individual subunits and subunit chimeras in transiently transfected COS-1 cells. Our results demonstrate that the H,K-ATPase alpha subunit requires its beta subunit for efficient cell-surface expression, as determined by indirect immunofluorescence. The H,K-ATPase beta protein appears to be able to get to the cell surface unaccompanied by any alpha subunit and appears to localize as well to a population of intracellular vesicles. We find that a transfected chimera encoding the NH2-terminal half of the H,K-ATPase alpha subunit and the COOH-terminal half of the Na,K-ATPase alpha subunit appears to assemble with the endogenous Na,K-ATPase beta subunit and to reach the plasmalemma. Transfection of the complementary alpha chimera requires coexpression with the H,K-ATPase beta subunit in order to attain surface delivery. Thus, it is the COOH-terminal half of the alpha subunit that specifies assembly with a particular beta subunit.
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PMID:Molecular requirements for the cell-surface expression of multisubunit ion-transporting ATPases. Identification of protein domains that participate in Na,K-ATPase and H,K-ATPase subunit assembly. 839 Sep 91

To investigate the regulation of the Na,K-ATPase, we have studied the expression of the Na,K-ATPase polypeptides in several mammalian cell lines using the vaccinia virus/T7 RNA polymerase expression system. Infection of several fibroblast-like cell lines with viral recombinants containing the Na,K-ATPase alpha and beta isoforms, the glucose transporters, GLUT 1 and GLUT 4, or the capsid protein of the Sindbis virus all result in the production of the appropriate protein products. However, all epithelial cell lines tested fail to synthesize the Na,K-ATPase viral recombinants, yet they efficiently express the other virally directed polypeptides. While Madin-Darby canine kidney (MDCK) epithelial cells infected with the Na,K-ATPase alpha1 or beta1 recombinant viruses produce both mRNAs, the messages are inefficiently translated. Furthermore, the RNA from infected MDCK cells does not direct the in vitro synthesis of the beta1 polypeptide, whereas the message from infected fibroblast-like BSC 40 cells is efficiently translated both in vivo and in vitro. Moreover, the synthesis of the H,K-ATPase alpha subunit is also limited in MDCK cells, although the H,K-ATPase beta subunit is efficiently expressed. Expression of chimeras constructed between the Na+ pump beta1 isoform and the H,K-ATPase beta subunit indicates that sequences in the 5' coding region of the beta1 message have an inhibitory effect; however, the stringent translational regulation of the beta1 isoform in MDCK cells requires the 5' and 3' regions of the coding sequence. The ability of the polarized cell lines to limit the synthesis of the Na+ pump polypeptides while expressing other vaccinia recombinants at high levels suggests that the polarized cells possess a stringent mechanism for the specific translational regulation of a select set of messages.
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PMID:Translational regulation of Na,K-ATPase alpha1 and beta1 polypeptide expression in epithelial cells. 879 17

The human ATP1AL1 gene belongs to the family of Na,K-ATPase and H,K-ATPase (X,K-ATPases) genes. It encodes a catalytic subunit of hitherto unknown human ouabain-sensitive H,K-ATPase that represents a novel third group of X,K-ATPases distinct from the known Na,K-ATPase and gastric H,K-ATPase. Cloning of the ATP1AL1 gene is described in this report. The exon-intron structure of ATP1AL1 was found to be very similar to that of related genes. It contains 23 exons and spans approximately 32 kb of genomic DNA. All ATP1AL1 exons and 12 of its 22 introns were entirely sequenced. A total of nine Alu repeats were identified in introns. The transcription initiation site was mapped 187 bp upstream of the ATG initiation codon by primer extension and S1 nuclease protection analyses of RNA from human skin and colon. Sequence analysis of the 5'-flanking region (1.48 kb) revealed numerous potential binding sites for transcription factors Sp1 and AP2 and one putative NF-kappa B binding site. The 0.85-kb region from position -484 (5'-flanking region) to position +369 (intron 1) meets the structural criteria of a CpG island. It is suggested that the ATP1AL1 gene contains two poly(A) addition sites that may function in a tissue-specific manner.
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PMID:Genomic organization of the human ATP1AL1 gene encoding a ouabain-sensitive H,K-ATPase. 883 94


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