Sodium and potassium ion-dependent change in oligomerization of Na,K-ATPase in C12E8 detected by low-angle laser light scattering technique in combination with high performance porous silica-gel chromatography.
Approximate molecular weights and the subunit structures of Na,K-ATPase from horse kidney were estimated by means of the combination of porous silica gel chromatography, laser light scattering (LS) and refractive index (RI) measurements in C12E8. When the enzymes were eluted with NaCl- or KCl-containing solution, 3 or 4 protein peaks, respectively were detected except that of low molecular weight range. These peaks were tentatively named Na-1, Na-2, Na-2', Na-3 (NaCl-containing eluents), K-1, K-2, K-3 (KCl-containing eluents), respectively. Na,K-ATPase and K-p-nitrophenylphosphatase activities were detected at all peaks. The activities were completely inhibited by ouabain. The ratios of the output from laser light scattering to that of differential refractive index intensity for reference proteins and these peaks were compared. Relative values of refractive index increments of BSA, thyroglobulin and C12E8 measured with the same RI detector under the same conditions were 0.144, 0.141, and 0.135 respectively. The size of the enzyme at the main peak (K-2) with K eluents (KCl 10 mM, 25 mM) was twice that at the main peak (Na-2') with Na eluents (1, 25, 50 mM NaCl) assuming that dn/dc of K-2 is similar to that of Na-2'. Na-3 and K-3 appeared at the same retention time and showed the same values of LS/RI. Provided that the dn/dc values of both peaks are similar to those of Na-2' and K-2, the sizes of Na-3 and K-3 are one-third of Na-2' and one-sixth of K-2, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006736:Horses; D007668:Kidney / Q000201:enzymology; D007834:Lasers; D046911:Macromolecular Substances; D008970:Molecular Weight; D011188:Potassium / Q000494:pharmacology; D012542:Scattering, Radiation; D012964:Sodium / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000302:isolation & purification / Q000378:metabolism*
D046911:Macromolecular Substances; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium