Buffer, pH, and ionic strength effects on the (Na+ + K+)-ATPase.
A dog kidney (Na+ + K+)-ATPase preparation also catalyzes K+-independent and K+-activated phosphatase reactions with p-nitrophenyl phosphate as substrate. K+-independent activity increases with declining pH over the range 7.5 to 5.8, whereas the other two activities decrease. The increased K+-independent activity is similar with imidazole, histidine, and several Good buffers, and is thus attributable to free H+, probably by affecting enzyme conformations rather than by changing affinity for Mg2+ or substrate or by H+ occupying specific K+-sites. The decrease in K+-phosphatase and (Na+ + K+)-ATPase activities with pH also occurs similarly with those buffers, and is not due to changes in apparent affinity for substrate or for cation activators. However, the Good buffers Pipes and ADA inhibit the K+-independent phosphatase reaction strongly, the K+-activated reaction moderately, and the (Na+ + K+)-ATPase reaction little; both contain two acidic groups, unlike the other buffers tested. Inhibition of the phosphatase reaction by Pipes is associated with a decreased apparent affinity for K+ and an increased sensitivity to inhibition by Na+ and ADP, consistent with Pipes hindering conformational transitions to the E2 enzyme forms required for phosphatase hydrolytic activity.
D000476:Alkanesulfonates; D017738:Alkanesulfonic Acids; D000818:Animals; D001667:Binding, Competitive; D002021:Buffers; D004285:Dogs; D006863:Hydrogen-Ion Concentration; D007098:Imino Acids; D007668:Kidney / Q000201:enzymology; D009596:Nitrophenols / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000378:metabolism; D009994:Osmolar Concentration; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D010879:Piperazines; D011188:Potassium / Q000494:pharmacology*; D011487:Protein Conformation; D011522:Protons; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D000476:Alkanesulfonates; D017738:Alkanesulfonic Acids; D002021:Buffers; D007098:Imino Acids; D009596:Nitrophenols; D009943:Organophosphorus Compounds; D010879:Piperazines; D011522:Protons; C008644:nitrophenylphosphate; C020184:N-(2-acetamido)iminodiacetic acid; D010744:Phosphoric Monoester Hydrolases; D000254:Sodium-Potassium-Exchanging ATPase; C008916:piperazine-N,N'-bis(2-ethanesulfonic acid); D011188:Potassium