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E Rabon

Publications and source records attributed to E Rabon.

28 records · Page 2Linked to original sources
PubMed1980

The interaction of K+ with gastric parietal cells and gastric ATPase.

The gastric H+ secretion in isolated cell requires K+ and is ATP dependent. There is also evidence in the cell system for Na+ inhibition of H+ secretion. The isolated gastric ATPase also shows K+ activation and inhibition by K+ or Na+ located on the ATP binding side of the enzyme, which corresponds to the cytoplasmic face of the enzyme. Gastric vesicles are activated in terms of transport activity by internal K+, and this site, inhibited by reagents that modify carboxyl groups, is required for enzyme turnover and transport.

Adenosine Triphosphate↗
PubMed1980

Aspects of parietal cell biology: cells and vesicles.

Many features of these gastric vesicles satisfy the requirements for the gastric H+ pump. For example, we have: (a) K+ requirement, (b) KA for K+ of about 30 mM; (c) identical cation sequence for tissue and vesicles, (d) similar anion sequence, (e) localization at the microvillus of the secretory canaliculus, (f) TI+ inhibiting H+ transport of both systems, and (g) the K+ gradient satisfying the osmotic gradient requirement for HCl-flow out of the parietal cell. Points that require explanation are lack of SCN- effects and regulation of KCl permeability.

Adenosine Triphosphatases↗
PubMed1979

Highly purified basal lateral plasma membranes from rat duodenum. Physical criteria for purity.

Preparations of intestinal epithelial cell basal lateral plasma membranes were analyzed with free flow electrophoresis and density perturbation with digitonin. The initial basal lateral membrane preparations were obtained by equilibrium density gradient centrifugation after two different schemes of homogenization and differential sedimentation (A.K. Mircheff, C.H. van Os, and E.M. Wright. 1978. Membr. Biochem. 1:177, and A.K. Mircheff, S.D. Hanna, M.W. Walling, and E.M. Wright. 1979. Prep. Biochem. 9:33. In these preparations, Na,K-ATPase, a marker for the basal lateral mambrane, was purified 16- to 18-fold over the initial homogenate. The preparations were also enriched in NADPH-cytochrome c reductase, alkaline phosphatase, acid phosphatase, and galactosyltransferase. Both free-flow electrophoresis, which separates on the basis of surface charge, and density perturbation with digitonin, which depends on a specific interaction of digitonin with cholesterol-rich membranes, resolved the preparation into three populations of particles. The major population, which represented basal lateral membranes purified 20- to 32-fold with respect to the initial homogenate, contained Na,K-ATPase, alkaline phosphatase, adenylate cyclase, and acid phosphatase. A second population was defined by its content of NADPH-cytochrome c reductase, and the third was defined by its content of galactosyltransferase. Guanylate cyclase appeared to be partitioned between the Na,K-ATPase-rich and NADPH-cytochrome c reductase-rich populations. Galactosyltransferase is also present in fractions which contain the Na,K-ATPase-rich membranes, but the present data cannot exclude the possibility of spillover by the adjacent, galactosyltransferase-rich population. This work emphasizes the importance of multiple, physical criteria for purity in the isolation of subcellular components.

Animals↗
PubMed1979

Induction of a chloride conductance in gastric vesicles by limited trypsin or chymotrypsin digestion or ageing.

Transport activity of the hog gastric (H+ + K+)-ATPase system was measured either as the formation of proton gradient using the dye probe acridine orange or as the formation of a proton diffusion potential using the cyanine dye 3,3'-diethyloxdicarbocyanine iodide in the presence of the protonophore tetrachlorosalicylanilide. The development of these gradients has been compared in K+ media in the presence of either Cl- or SO4-2 as the anionic species. This comparison of proton diffusion potential formation to proton gradient formation has been used to demonstrate that a Cl- conductance in this vesicular system results from limited enzymic digestion with either trypsin or alpha-chymotrypsin from the ageing process itself. The possible significance of this finding is discussed.

Acridine Orange↗
PubMed1979

Transport characteristics of frog gastric membranes.

ATP-induced transport by fractions of frog gastric microsomes prepared either by density gradient centrifugation of by free flow electrophoresis were K+ dependent and hence considered due to a K+-activated ATPase. Significant activity of this enzyme was, however, only found in the anodic peak of the free flow electrophoretic separation, which in addition to separating transporting from non-transporting particles, also separated membranes containing a phosphorylatable peptide (Mr=105 000) region as the major peptide on SDS-polyacrylamide gel electrophoresis from those containing a peptide (Mr=44 000) on SDS-polyacrylamide gel electrophoresis. H+ uptake, measured either by acridine orange or 3,3'-diethyloxadicarbocyanine + tetrachlorosalicylanilide absorbance changes was dependent on K+ intravesicularly. Using 86Rb+, active extrusion of the cation followed ATP addition. SCN-, an inhibitor of acid secretion did not affect the latter, but blocked signals due to H+ uptake, in contrast to mammalian preparations.

Adenosine Triphosphatases↗
PubMed1978

Quantitation of hydrogen ion and potential gradients in gastric plasma membrane vesicles.

The ATP-dependent uptake of H+ by hog gastric parietal cell vesicles was quantitated by using the pH indicator dyes bromcresol green and malachite green, the weak bases, aminopyrine and 9-aminoacridine, and the pH electrode. A K+-dependent H+ uptake was found, with a significant difference between the quantity of H+ disappearing from the medium (deltaHo) and the quantity appearing inside the vesicle (deltaHi). 9-Aminoacridine gave a lower value for the deltaHi than any of the other probes. Probes of potential such as diethyloxadicarbocyanine or oxonol dyes showed that only secondary diffusion potentials occurred during H+ uptake and that the cationic dyes in the presence of protonophores could also be used to quantitate H+ uptake. The potential in the presence of protonophore indicated a deltaHi greater than that found with the other probes. Binding sites for acridine orange were generated either by ATP or an artificial pH gradient and corresponded to the deltaHi indicated by aminopyrine. SCN- (30mM) only partially inhibited the H+ gradient, and this, coupled with the failure to detect the physiological deltapH of 6.6, indicated that these vesicles may be an incomplete model of gastric acid secretion.

Adenosine Triphosphate↗
PubMed1977

Proton transport by gastric membrane vesicles.

A highly purified membrane fraction was derived from hog gastric mucosa by a combination of differential and density gradient centrifugation and free flow electrophoresis. This final fraction was 35-fold enriched with respect to cation activated ouabain-insensitive ATPase. Antibody against this fraction was shown to be bound to the luminal surface of the gastric glands. The addition of ATP to this fraction or the density gradient fraction resulted in H+ uptake into an osmotically sensitive space. The apparent Km for ATP was 1.7-10(-4) M in the absence of a K+ gradient similar to that found for ATPase activity. The reaction is specific for ATP and requires cation in the sequence K+ greater than Rb+ greater than Cs+ greater than Na+ greater than Li+ and inhibited by ATPase inhibitors such as N,N'-dicylclohexyl-carbodiimide. Maximal H+ uptake occurs with an outward K+ gradient but the minimal apparent KA is found in the absence of a K+ gradient. The pH optimum for H+ uptake is between 5.8 and 6.2 which corresponds to the pH range for phosphroylation of the enzyme, but is considerably less than the pH maximum of the K+ dependent dephosphorylation. In the presence of an inward K+ gradient, protonophores such as tetrachlorsalicylanilide only partially abolish the H+ gradient but valinomycin dissipates 75% of the gradient, and nigericin abolishes the gradient. The vesicles therefore have a low K+ conductance but a measurable H+ conductance, hence a K+ gradient can produce an H+ gradient in the presence of valinomycin. The uptake and spontaneous leak of H+ are temperature sensitive with a similar transition temperature. Ultraviolet irradiation inactivates ATPase and proton transport at the same rate, approximately at twice the rate of p-nitrophenylphosphatase inactivation. It is concluded that H+ uptake by these vesicles is probably due to a dimeric (H+ + K+)-ATPase and is probably non-electrogenic.

Adenosine Triphosphatases↗
PubMed1977

Metabolic and membrane aspects of gastric H+ transport.

Metabolic properties of dog gastric mucosa, investigated by substrate level measurements, implicate the Krebs cycle as the major energy-yielding metabolic pathway but are equivocal in terms of an ATP-based H+ secretion. Purification of gastric membranes by centrifugation and free flow electrophoresis results in a class of membrane vesicles enriched in K+-ATPase and capable of ATP-energized H+ uptake. Immunohistochemistry shows these to be derived from the parietal cell. H+ uptake by the vesicles is accompanied by K+ efflux, and movement of either ion is not potential-coupled. The simplest interpretation of these transport studies is uptake of KCl by the vesicles by passive diffusion followed by active H+:K+ exchange. In some respects, however, this model fails to conform to the expectations from in vitro studies. It may be, therefore, that another pump (i.e., redox) or another membrane component (i.e., Cl- conductance) is lost during purification. The properties of the vesicles are such, however, as to establish their role in H+ secretion by the stomach.

Adenosine Diphosphate↗
PubMed1976

A nonelectrogenic H+ pump in plasma membranes of hog stomach.

Differential and density gradient centrifugation were used to prepare a vesicular membrane fraction from hog gastric mucosa enriched 17-fold with respect to cation-activated ATPase and 5'-AMPase. Fractionation of the gradient material by free flow electrophoresis resulted in a fraction 35-fold enriched in cation-activated ATPase and essentially free of 5'-AMPase and Mg2+ATPase. The addition of ATP to either fraction resulted in H+ uptake and Rb+ efflux. The ionophoric and osmotic sensitivity showed that these ion movements were due to transport rather than binding. The cation selectivity sequences, substrate specificities and action of inhibitors indicated that the transport was a function of K+ATPase activity. The characteristics of the ATP-dependent enhancement of SCN- uptake and 8-anilinonapthalene-1-sulfonate fluorescence in the presence of valinomycin and the action of ionophores and lipid-permeable ions suggested that the energy dependent K+:H+ exchange was effectively nonelectrogenic. Thus these vesicles contain a nonelectrogenic (H+ + K+)-ATPase, hence acid secretion by the stomach is probably due to an ATP-dependent H+ + K+ exchange.

Adenosine Triphosphatases↗