Summary
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1.
The distribution of ATPase and several marker enzymes was examined after differential and sucrose gradient centrifugation of yeast homogenates.
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2.
An ATPase activity not sensitive to oligomycin is found exclusively associated with a particulate fraction equilibrating at densities of 1.23–1.25. This particulate material shows the chemical and enzymatic characteristics of the yeast plasma membrane.
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3.
The pH optimum of the plasma membrane ATPase is 5.6, as compared with 8.5 for the mitochondrial ATPase. In addition to oligomycin, the enzyme is not sensitive to other inhibitors of the mitochondria) ATPase as azide, dicyclohexylcarbodiimide and the mitochondrial ATPase inhibitor protein. It is inhibited by p-chloromercuryphenyl sulfonate, fluoride, quercetin and by the antibiotic Dio-9 but is not affected by ouabain.
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4.
The plasma membrane ATPase shows a high affinity for ATP (Km=0.1 mm) and is very specific for this compound, hydrolyzing other nucleotide triphosphates less than 25% as rapidly. No activity was detected with ADP.
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5.
The enzyme requires a divalent cation for activity and Mg2+ is the most effective. It is not significantly stimulated by K+ or bicarbonate and Ca2+ is inhibitory.
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6.
The activity cannot be assayed in intact cells unless they are permeabilized with toluene. This suggest that the active site is on the cytoplasmic side of the plasma membrane.
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Serrano, R. Characterization of the plasma membrane ATPase of Saccharomyces cerevisiae. Mol Cell Biochem 22, 51–63 (1978). https://doi.org/10.1007/BF00241470
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DOI: https://doi.org/10.1007/BF00241470