Na+ATPase of the mammalian erythrocyte membrane. Reversibility of phosphorylation at 0 degrees.
When human erythrocyte membranes are phosphorylated with a very low concentration of [gamma-32P]ATP (0.02 muM) at 0 degrees, and then EDTA is added, rapid disappearance of the phosphoenzyme intermediate of Na+ATPase is observed. The initial rapid phase of phosphoenzyme disappearance is, for the most part, not associated with P1 release and its rate constant, kD, is severalfold greater than the ratio of Na+ATPase activity to phosphoenzyme intermediate, v:EP, at steady state. It is concluded that this rapid disappearance of phosphoenzyme is due to resynthesis of ATP via reversal of phosphorylation. In contrast, rapid reversal is not observed when excess nonradioactive ATP is added to reduce E32P formation, provided Mg2+ is present; however, K+ added with the ATP stimulates reversal. Rapid reversal following EDTA addition is unlikely also when higher ATP concentrations (greater than or equal to 10(-6) M) are used to phosphorylate the enzyme since, at higher ATP, kD congruent to v:EP. The results are compatible with the concept that the Na+ATPase enzyme is composed of two or more catalytic subunits, in which ATP at one catalytic site can regulate the reactivity at another site.
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000251:Adenosine Triphosphatases / Q000097:blood*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D004492:Edetic Acid / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004912:Erythrocytes / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009840:Oligomycins / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology*; D013696:Temperature; D013997:Time Factors
D009840:Oligomycins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D012964:Sodium; D000251:Adenosine Triphosphatases; D008274:Magnesium; D011188:Potassium