Characterization of the phosphorylated intermediate of the K+-translocating Kdp-ATPase from Escherichia coli. 1989

A Siebers, and K Altendorf
Arbeitsgruppe Mikrobiologie, Universität Osnabrück, Federal Republic of Germany.

During ATP hydrolysis the K+-translocating Kdp-ATPase from Escherichia coli forms a phosphorylated intermediate as part of the catalytic cycle. The influence of effectors (K+, Na+, Mg2+, ATP, ADP) and inhibitors (vanadate, N-ethylmaleimide, bafilomycin A1) on the phosphointermediate level and on the ATPase activity was analyzed in purified wild-type enzyme (apparent Km = 10 microM) and a KdpA mutant ATPase exhibiting a lower affinity for K+ (Km = 6 mM). Based on these data we propose a minimum reaction scheme consisting of (i) a Mg2+-dependent protein kinase, (ii) a Mg2+-dependent and K+-stimulated phosphoprotein phosphatase, and (iii) a K+-independent basal phosphoprotein phosphatase. The findings of a K+-uncoupled basal activity, inhibition by high K+ concentrations, lower ATP saturation values for the phosphorylation than for the overall ATPase reaction, and presumed reversibility of the phosphoprotein formation by excess ADP indicated similarities in fundamental principles of the reaction cycle between the Kdp-ATPase and eukaryotic E1E2-ATPases. The phosphoprotein was tentatively characterized as an acylphosphate on the basis of its alkali-lability and its sensitivity to hydroxylamine. The KdpB polypeptide was identified as the phosphorylated subunit after electrophoretic separation at pH 2.4, 4 degrees C of cytoplasmic membranes or of purified ATPase labeled with [gamma-32P]ATP.

UI MeSH Term Description Entries
D007700 Kinetics The rate dynamics in chemical or physical systems.
D008565 Membrane Proteins Proteins which are found in membranes including cellular and intracellular membranes. They consist of two types, peripheral and integral proteins. They include most membrane-associated enzymes, antigenic proteins, transport proteins, and drug, hormone, and lectin receptors. Cell Membrane Protein,Cell Membrane Proteins,Cell Surface Protein,Cell Surface Proteins,Integral Membrane Proteins,Membrane-Associated Protein,Surface Protein,Surface Proteins,Integral Membrane Protein,Membrane Protein,Membrane-Associated Proteins,Membrane Associated Protein,Membrane Associated Proteins,Membrane Protein, Cell,Membrane Protein, Integral,Membrane Proteins, Integral,Protein, Cell Membrane,Protein, Cell Surface,Protein, Integral Membrane,Protein, Membrane,Protein, Membrane-Associated,Protein, Surface,Proteins, Cell Membrane,Proteins, Cell Surface,Proteins, Integral Membrane,Proteins, Membrane,Proteins, Membrane-Associated,Proteins, Surface,Surface Protein, Cell
D008970 Molecular Weight The sum of the weight of all the atoms in a molecule. Molecular Weights,Weight, Molecular,Weights, Molecular
D010750 Phosphoproteins Phosphoprotein
D010766 Phosphorylation The introduction of a phosphoryl group into a compound through the formation of an ester bond between the compound and a phosphorus moiety. Phosphorylations
D011188 Potassium An element in the alkali group of metals with an atomic symbol K, atomic number 19, and atomic weight 39.10. It is the chief cation in the intracellular fluid of muscle and other cells. Potassium ion is a strong electrolyte that plays a significant role in the regulation of fluid volume and maintenance of the WATER-ELECTROLYTE BALANCE.
D004492 Edetic Acid A chelating agent that sequesters a variety of polyvalent cations such as CALCIUM. It is used in pharmaceutical manufacturing and as a food additive. EDTA,Edathamil,Edetates,Ethylenediaminetetraacetic Acid,Tetracemate,Calcium Disodium Edetate,Calcium Disodium Versenate,Calcium Tetacine,Chelaton 3,Chromium EDTA,Copper EDTA,Coprin,Dicobalt EDTA,Disodium Calcitetracemate,Disodium EDTA,Disodium Ethylene Dinitrilotetraacetate,Distannous EDTA,Edetate Disodium Calcium,Edetic Acid, Calcium Salt,Edetic Acid, Calcium, Sodium Salt,Edetic Acid, Chromium Salt,Edetic Acid, Dipotassium Salt,Edetic Acid, Disodium Salt,Edetic Acid, Disodium Salt, Dihydrate,Edetic Acid, Disodium, Magnesium Salt,Edetic Acid, Disodium, Monopotassium Salt,Edetic Acid, Magnesium Salt,Edetic Acid, Monopotassium Salt,Edetic Acid, Monosodium Salt,Edetic Acid, Potassium Salt,Edetic Acid, Sodium Salt,Ethylene Dinitrilotetraacetate,Ethylenedinitrilotetraacetic Acid,Gallium EDTA,Magnesium Disodium EDTA,N,N'-1,2-Ethanediylbis(N-(carboxymethyl)glycine),Potassium EDTA,Stannous EDTA,Versenate,Versene,Acid, Edetic,Acid, Ethylenediaminetetraacetic,Acid, Ethylenedinitrilotetraacetic,Calcitetracemate, Disodium,Dinitrilotetraacetate, Disodium Ethylene,Dinitrilotetraacetate, Ethylene,Disodium Versenate, Calcium,EDTA, Chromium,EDTA, Copper,EDTA, Dicobalt,EDTA, Disodium,EDTA, Distannous,EDTA, Gallium,EDTA, Magnesium Disodium,EDTA, Potassium,EDTA, Stannous,Edetate, Calcium Disodium,Ethylene Dinitrilotetraacetate, Disodium,Tetacine, Calcium,Versenate, Calcium Disodium
D004926 Escherichia coli A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc. Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D006863 Hydrogen-Ion Concentration The normality of a solution with respect to HYDROGEN ions; H+. It is related to acidity measurements in most cases by pH pH,Concentration, Hydrogen-Ion,Concentrations, Hydrogen-Ion,Hydrogen Ion Concentration,Hydrogen-Ion Concentrations
D000227 Adenine Nucleotides Adenine Nucleotide,Adenosine Phosphate,Adenosine Phosphates,Nucleotide, Adenine,Nucleotides, Adenine,Phosphate, Adenosine,Phosphates, Adenosine

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