Modified cation activation of the (Na+K)-ATPase following treatment with thimerosal.
Treatment of the Na,K-ATPase enzyme, isolated from canine renal outer medulla, with thimerosal (ethylmercurithiosalicylate) resulted in significant inhibition of the overall Na,K-ATPase with only slight, if any, inhibition of the Na-ATPase and ATP:ADP exchange activities. The K-stimulated PNPPase activity was stimulated [see G. R. Henderson and A. Askari (1977) Arch. Biochem. Biophys. 182, 221-226]. Examination of the Na dependence of the ATPase and ATP:ADP exchange activities revealed an Na-independent, ouabain-sensitive activity that was inhibited by Na in the range 0-10 mM. At greater than or equal to 10 mM concentration of Na the treated and modified enzymes showed similar activities. The apparent affinity of the modified enzyme for ATP in the presence of 100 mM Na was the same as that of the untreated enzyme (0.2-0.3 microM). In the absence of Na, the modified enzyme hydrolyzed ATP with a relatively low affinity (about 120 microM). The enhancement of p-nitrophenylphosphatase (PNPPase) activity measured in the presence of K ions was due to the appearance of K-independent, ouabain-sensitive PNPP activity. The modification was without major affect on the apparent affinity of the enzyme for K ions in the PNPPase activity. Treatment of the thimerosal-modified enzyme with dithiothreitol removed (or greatly reduced) the cation-independent, ouabain-sensitive activities and the Na,K-ATPase activity returned. Modification of a set of enzyme -SH groups in the Na,K-ATPase enzyme made it able to hydrolyze ATP in the absence of Na ions and PNPP in the absence of K ions. The -SH groups modified by thimerosal are evidently critical to the major dephosphoenzyme conformational changes but are not involved in the major transport conformational change between phosphoenzymes.
Kaplan
J H
JH
Mone
M D
MD
D009597:4-Nitrophenylphosphatase / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002412:Cations / Q000494:pharmacology*; D004285:Dogs; D004789:Enzyme Activation / Q000187:drug effects; D005035:Ethylmercury Compounds / Q000494:pharmacology*; D066298:In Vitro Techniques; D007668:Kidney / Q000201:enzymology; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013849:Thimerosal / Q000494:pharmacology*
D002412:Cations; D005035:Ethylmercury Compounds; D013849:Thimerosal; D010042:Ouabain; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D012964:Sodium; D009597:4-Nitrophenylphosphatase; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium