Selective solubilization with tween 20 of proteins from water-extracted human erythrocyte membranes analysis by gel electrophoresis in dodecylsulfate and in tween 20

Abstract

1. 1. Human erythrocyte membranes (ghosts) were extracted at pH 9.5 (0°C) with an aqueous solution of low ionic strength (containing EDTA and 2-mercaptoethanol) and then with water. This procedure selectively solubilized 35% of the membrane protein. 2. 2. The material not solubilized by the above procedure (the membrane residue) was partially solubilized with the non-ionic detergent Tween 20 at an ionic strength of approx. 0.005. The degree of solubilization increased with increasing pH (4.5–9.8) and with increasing detergent concentration. 3. 3. The solubilization of proteins with Tween 20 at pH 9.8 was highly selective at an ionic strength of approx. 0.05 as well as at low concentrations of detergent at an ionic strength of approx. 0.005, as indicated by polyacrylamide-gel electrophoresis in dodecylsulfate, only the two major components of high molecular weight (1,2) and one smaller component (4.1) being solubilized. 4. 4. Membrane residue protein solubilized with Tween 20 separated into 5–6 major zones upon electrophoresis in a polyacrylamide gel containing 0.5% Tween 20 and 0.02 M glycine-NaOH buffer (pH 9.8), as seen after staining with Coomassie Brilliant Blue. 5. 5. The patterns observed in two-dimensional polyacrylamide-gel electrophoresis using Tween 20 in the first and dodecylsulfate in the second direction suggest that some of the zones obtained by electrophoresis in Tween 20 contained protein complexes which were split by dodecylsulfate.