Ouabain-binding site of (Na+ + K+)-ATPase in right-side-out vesicles has not an externally accessible SH group.
The fluorescing sulfhydryl reagent N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM) inactivates purified (Na+ + K+)-ATPase at 20 microM. This inactivation results in a decrease of the ouabain-binding capacity of the enzyme. Treatment of (Na+ + K+)-ATPase, embedded in right-side-out-oriented vesicles, by DACM does not affect ouabain binding to the enzyme. Incorporation of DACM into the alpha subunit of (Na+ + K+)-ATPase embedded in right-side-out vesicles is also not affected by the presence or absence of 100 microM ouabain. It is therefore concluded that a sulfhydryl group does not reside within the ouabain-binding site of (Na+ + K+)-ATPase.
D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D007679:Kidney Medulla / Q000201:enzymology; D008301:Maleimides / Q000494:pharmacology; D008565:Membrane Proteins / Q000378:metabolism; D010042:Ouabain / Q000378:metabolism*; D011955:Receptors, Drug / Q000378:metabolism*; D012756:Sheep; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D008301:Maleimides; D008565:Membrane Proteins; D011955:Receptors, Drug; D013438:Sulfhydryl Compounds; D013439:Sulfhydryl Reagents; C028996:cardiac glycoside receptors; C019347:N-(7-dimethylamino-4-methylcoumarinyl)maleimide; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase