Phase separation of nonionic detergents by salt addition and its application to membrane proteins.
By adding salts (sodium chloride, ammonium sulfate), it is possible to induce phase separation in membrane-protein solubilisates containing Triton X-100 or Nonidet P-40 at temperatures between 0 and 20 degrees C. Other nonionic detergents of the Brij, Lubrol, and Tween series can also be used for this procedure. The salt concentration required for induction of phase separation is dependent on the hydrophobicity of the detergent used. For detergents of the Triton series it seems that detergents with lower hydrophilic lipophilic balance numbers need lower salt concentrations to separate the phases than those with larger hydrophilic parts. Ammonium sulfate precipitation as an initial purification step for membrane proteins should be avoided in the presence of the nonionic detergents tested. Instead of this procedure, phase separation induced by sodium chloride or ammonium sulfate can be recommended, as was proved for membrane-bound proteases of Pseudomonas aeruginosa and Bacillus cereus and for bacteriorhodopsin.
Fricke
B
B
Department of Medicine, Martin Luther University, Halle, Germany.
D000645:Ammonium Sulfate; D001426:Bacterial Proteins / Q000302:isolation & purification; D001436:Bacteriorhodopsins / Q000302:isolation & purification; D003902:Detergents / Q000302:isolation & purification*; D010450:Endopeptidases / Q000302:isolation & purification; D005069:Evaluation Studies as Topic; D008565:Membrane Proteins / Q000302:isolation & purification*; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000302:isolation & purification; D012965:Sodium Chloride
D001426:Bacterial Proteins; D003902:Detergents; D008565:Membrane Proteins; D011092:Polyethylene Glycols; D012965:Sodium Chloride; D001436:Bacteriorhodopsins; D017830:Octoxynol; C010615:Nonidet P-40; D010450:Endopeptidases; D000645:Ammonium Sulfate