Phenyl-Sepharose-mediated detergent-exchange chromatography: its application to exchange of detergents bound to membrane proteins.
Detergent-saturated phenyl-Sepharose was used to exchange detergents for one another in the presence of membrane proteins. The alkyl detergents lauryl maltoside, octyl glucoside, and dodecyl sulfate were each successfully exchanged for Triton X-100, Triton N-101, or Nonidet P-40 present in a solution of either cytochrome c oxidase, a mixture of inner mitochondrial membrane proteins, or a mixture of erythrocyte membrane proteins. The method involves (1) saturating a small column of phenyl-Sepharose (1-2 mL) with one of the alkyl detergents at a pH of 8 or 9 and an ionic strength of 0.01, (2) applying a detergent-solubilized membrane protein sample containing as much as 20 mg/mL of Triton X-100, Triton N-101, or Nonidet P-40, and (3) eluting the protein with buffer containing the detergent with which the resin had been saturated. With this approach, 90-99% of the detergent in the initial protein sample was exchanged for the second detergent with an 80-100% recovery of protein. The advantages of this method over previous approaches for exchanging detergents include the rapidity of the technique and the apparent general applicability of the method to a wide variety of detergents and membrane proteins.
D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity; D003902:Detergents; D003576:Electron Transport Complex IV; D005233:Fatty Alcohols; D005960:Glucosides; D006801:Humans; D008565:Membrane Proteins / Q000302:isolation & purification*; D017137:Nonoxynol; D017830:Octoxynol; D011092:Polyethylene Glycols; D012441:Saccharomyces cerevisiae; D012685:Sepharose / Q000031:analogs & derivatives; D012967:Sodium Dodecyl Sulfate; D013501:Surface-Active Agents
D003902:Detergents; D005233:Fatty Alcohols; D005960:Glucosides; D008565:Membrane Proteins; D013501:Surface-Active Agents; D017137:Nonoxynol; C018619:octyl-beta-D-glucoside; D012967:Sodium Dodecyl Sulfate; D011092:Polyethylene Glycols; C020927:phenyl-sepharose; D017830:Octoxynol; D012685:Sepharose; C010615:Nonidet P-40; C028913:dodecyl sulfate; D003576:Electron Transport Complex IV