Phenyl-Sepharose chromatography of membrane proteins solubilized in Triton X-100

Chromatography on phenyl-Sepharose provides a convenient means of concentrating "membrane" proteins, which bind Triton X-100, and of separating them from "soluble" proteins, which do not bind the detergent. As an example, chromatography on phenyl-Sepharose has been used in the purification of tissue factor from human brain and placenta. Triton X-100 extracts of brain and placenta are separated into two distinct protein fractions when chromatographed on phenyl-Sepharose. One fraction elutes in detergent-free buffer, whereas the other elutes only after saturation of the column with Triton X-100. The absence of proteins in fractions eluted prior to detergent saturation of the column suggested that, in the presence of Triton X-100, protein binding to phenyl-Sepharose occurred as a complex with the detergent. A propylene glycol concentration gradient eluted bound proteins and Triton X-100 over the same concentrations of propylene glycol.