Fractionation of membrane proteins by temperature-induced phase separation in Triton X-114. Application to subcellular fractions of the adrenal medulla.
After solubilization with the detergent Triton X-114, membrane proteins may be separated into three groups: if the membrane is sufficiently lipid-rich, one family of hydrophobic constituents separates spontaneously at low temperature; warming at 30 degrees C leads to separation of a detergent-rich phase and an aqueous phase. Using the chromaffin-granule membrane as a model, we found that many intrinsic membrane glycoproteins are found in the latter phase, probably maintained in solution by adherent detergent. They precipitate, however, when this is removed by dialysis, leaving in solution those truly hydrophilic proteins that were originally adhering to the membranes. We have used this method with mitochondria, and with Golgi- and rough-endoplasmic-reticulum-enriched microsomal fractions: it has proved to be a rapid and convenient method for effecting a partial separation of proteins from a variety of different membranes.
D000251:Adenosine Triphosphatases / Q000302:isolation & purification; D000313:Adrenal Medulla / Q000032:analysis; D000818:Animals; D002417:Cattle; D002837:Chromaffin Granules / Q000032:analysis; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000302:isolation & purification; D008565:Membrane Proteins; D008722:Methods; D017830:Octoxynol; D011092:Polyethylene Glycols; D012995:Solubility; D013347:Subcellular Fractions / Q000032:analysis; D013501:Surface-Active Agents; D013696:Temperature
D003902:Detergents; D006023:Glycoproteins; D008565:Membrane Proteins; D013501:Surface-Active Agents; D011092:Polyethylene Glycols; D017830:Octoxynol; C010615:Nonidet P-40; D000251:Adenosine Triphosphatases