SOLUBILIZATION AND MOLECULAR-WEIGHT DETERMINATION OF THE (NA++K+)-ATPASE FROM RECTAL GLANDS OF SQUALUS-ACANTHIAS

The membrane-bound (Na+ +K+)-activated ATPase (ATP Phosphohydrolase, EC 3.6.1.3) system was treated with the nonionic detergent octaethylene-glycoldodecyl ether, yielding a transparent supernatant after centrifugation. The supernatant was highly active with both ATPase and p-nitrophenylphosphatase, with initial specific activities of 2300 μmol Pi released · mg-1 protein · h-1 and 350 μmol p-nitrophenol released · mg-1 protein · h-1, respectively. The supernatant was purified to 95-100%, with respect to the 96 000 dalton and the 56 000 dalton peptides. The solubilized enzyme was gel filtered in Sepharose 4B-CL and displayed 2 peaks, both with catalytic activity. The low molecular weight particles eluted at Kav = 0.54, corresponding to a molecular weight of approximately 500 000 daltons and the particles had a specific activity of 2100 μmol Pi · mg-1 protein · h-1. Both peaks contained phospholipid with 60 mol phospholipid bound per 300 000 g protein. The low molecular weight particles had a molecular weight of 276 000 as determined by sedimentation equilibrium analysis. © 1979.