SEPARATION OF MEMBRANE-PROTEINS SOLUBILIZED WITH A NONDENATURING DETERGENT AND A HIGH-SALT CONCENTRATION BY HYDROXYAPATITE HIGH-PERFORMANCELIQUID-CHROMATOGRAPHY

Ceramic hydroxyapatite high-performance liquid chromatography with a s olvent containing high concentrations of salts was used for the separa tion of membrane proteins solubilized under nondenaturing and high sal t conditions. The chromatographic conditions were optimized using sodi um cholate as the detergent. By this method, most membrane proteins, p repared from rat liver rough microsomes, were effectively resolved fro m each other with a protein recovery of more than 90%. The method also allowed the single-step purification of the ribosome-binding protein, p34, from a microsomal membrane protein fraction. The good resolution with this method should be applicable to the isolation and characteri zation of a variety of membrane proteins on the analytical and semipre parative scales. With only the substitution of sodium cholate with oth er nondenaturing detergents, this method may also be applicable to the purification of membrane proteins requiring such nondenaturing deterg ents with retention of their biological activities. (C) 1995 Academic Press, Inc.