Duality of alpha-subunit of canine kidney sodium- and potassium-activated adenosinetriphophatase in electrophoresis.
Sodium- and potassium-activated adenosinetriphosphatase (Na+, K+-ATPase) purified from dog kidney outer medulla was examined by polyacrylamide gel electrophoresis and by photoaffinity labeling with N-(ouabain)-N'-(2-nitro-4-azidophenyl)-ethylenediamine (NAP-ouabain). The large subunit band (alpha-band) split into two bands on the gel after the enzyme was heat-treated in the presence of 1% sodium dodecylsulfate (SDS). Of the two bands (alpha I and alpha II), alpha I had the same electrophoretic mobility as the original band, while alpha II moved slightly faster. Total conversion into alpha II was not observed, about half of the original remaining as alpha I. Below 60 degree C, heat treatment did not produce alpha II. Phenylmethylsulfonyl fluoride did not prevent the appearance of alpha II. Both alpha I and alpha II were labeled with [3H]NAP-ouabain. Nonspecific incorporation of [3H]NAP-ouabain also occurred irrespective of illumination, but it was removed either by diffusion during staining and destaining of the gel or by treatment of the enzyme with trichloroacetic acid. It is tentatively concluded that the splitting of the band reflects some intrinsic differences in situ of the alpha-subunit of dog kidney membrane Na+,K+-ATPase.
D000345:Affinity Labels; D000818:Animals; D001386:Azides / Q000528:radiation effects; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D007679:Kidney Medulla / Q000201:enzymology*; D007700:Kinetics; D008027:Light; D008861:Microsomes / Q000201:enzymology; D010042:Ouabain / Q000031:analogs & derivatives / Q000528:radiation effects; D000254:Sodium-Potassium-Exchanging ATPase / Q000302:isolation & purification*; D014316:Tritium
D000345:Affinity Labels; D001386:Azides; D014316:Tritium; D010042:Ouabain; C032780:NAP-ouabain; D000254:Sodium-Potassium-Exchanging ATPase