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 mumol Pi released . mg-1 protein. h-1 and 350 mumol 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 mumol 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.
D009597:4-Nitrophenylphosphatase / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D003902:Detergents / Q000494:pharmacology; D004284:Dogfish; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D010455:Peptides / Q000032:analysis; D010743:Phospholipids / Q000032:analysis; D012007:Rectum / Q000201:enzymology; D012491:Salt Gland / Q000201:enzymology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000302:isolation & purification / Q000378:metabolism*; D012995:Solubility; D013696:Temperature
D003902:Detergents; D010455:Peptides; D010743:Phospholipids; D009597:4-Nitrophenylphosphatase; D000254:Sodium-Potassium-Exchanging ATPase