An essential arginine residue in the ATP-binding centre of (Na+ + K+)-ATPase.
1. Incubation of purified (Na+ + K+)-ATPase (ATP phosphohydrolase EC 3.6.1.3) from rabbit kidney outer medulla with butanedione in borate buffer leads to reversible inactivation of the (Na+ + K+)-ATPase activity. 2. The reaction shows second-outer kinetics, suggesting that modification of a single amino acid residue is involved in the inactivation of the enzyme. 3. The pH dependence of the reaction and the effect of borate ions strongly suggest that modification of an arginine residue is involved. 4. Replacement of Na+ by K+ in the butanedione medium decreases inactivation. 5. ATP, ADP and adenylyl imido diphosphate, particularly in the presence of trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid to complex Mg2+, protect the enzyme very efficiently against inactivation by butanedione. 6. The (Na+ + Mg2+)-dependent phosphorylation capacity of the enzyme is inhibited in the same degree as the (Na+ + K+)-ATPase activity by butanedione. 7. The K+-stimulated p-nitrophenylphosphatase activity is much less inhibited than the (Na+ + K+)ATPase activity. 8. The ATP stimulation of the K+-stimulated p-nitrophenylphosphatase activity is inhibited by butanedione to the same extent as the (Na+ + K+)-ATPase activity. 9. Modification of sulfhydryl groups with 5,5'-dithiobis(2-nitrobenzoic acid) protects partially against the inactivating effect of butanedione. 10. The results suggest that an arginine residue is present in the nucleotide binding centre of the enzyme.
D009597:4-Nitrophenylphosphatase / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate; D000818:Animals; D001120:Arginine / Q000032:analysis; D001665:Binding Sites; D003931:Diacetyl / Q000494:pharmacology; D004789:Enzyme Activation; D007679:Kidney Medulla / Q000201:enzymology; D007700:Kinetics; D011188:Potassium / Q000494:pharmacology; D011485:Protein Binding; D011817:Rabbits; D012964:Sodium / Q000494:pharmacology
D000255:Adenosine Triphosphate; D001120:Arginine; D012964:Sodium; D009597:4-Nitrophenylphosphatase; D000251:Adenosine Triphosphatases; D003931:Diacetyl; D011188:Potassium