High-performance liquid chromatographic separation of membrane proteins isolated from erythrocyte ghosts

Abstract

Membrane proteins extracted from erythrocyte ghosts with sodium dodecyl sulphate (SDS), 3-(3-cholamidopropyl)-dimethylamminopropane sulfonate (CHAPS) or octylglucoside have been analyzed in various reversed-phase high-performance liquid chromatographic systems. Only SDS was able to solubilize considerable amounts of membrane proteins with mol.wt. > 15 000 daltons, but these membrane proteins were recovered in poor yield from a silica-based C 4 column eluted with an acetonitrile gradient in trifluoroacetic acid (TFA). A resin-based phenyl column eluted with a similar TFA-acetonitrile gradient was found to be a better choice with respect to the recovery of membrane proteins with mol.wt. > 15 000 daltons, and when this column was eluted with an acetic acid gradient with increasing amounts of acetonitrile, erythrocyte ghost membrane proteins solubilized in SDS (mol.wt. 10 000–200 000 daltons) were separated in six major and several minor components with satisfactory recovery.