Phenyl-Sepharose-mediated detergent-exchange chromatography: its application to exchange of detergents bound to membrane proteins

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Abstract
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 [yeast] inner mitochondrial proteins, or a mixture of [human] erythrocyte membrane proteins. The method involves saturating a small column of phenyl-Sepharose (1-2 m/l) with one of the alkyl detergents at a pH of 8 or 9 and an ionic strength of 0.01, 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 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 2nd 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.