Association of protein fractions and lipids from human erythrocyte membranes. I. Studies on a strongly bound protein fraction

Half of the protein content of human erythrocyte membranes was removed from the membrane by 10% acetic acid. The remaining lipoprotein complex was depolymerized by 90%acetic acid. Protein and lipid were separated by gel filtration, and the protein fraction was transferred into aqueous buffers. Association in aqueous buffers could be observed between the protein fraction and sonicated total lipids from human erythrocyte membranes. Lipoprotein formation was possible over a wide range of conditions with respect to pH and ionic strength, including physiological conditions, and to protein and lipid concentrations. After equilibrium centrifugation in a density gradient, the lipoproteins formed clearly visible, narrow bands which contained up to 70% of the proteins and the lipids applied to the gradient. The density of the bands did not depend significantly on pH, ionic strength or the kind of ions present during recombination. It was, however, strongly influenced by the weight ratio of protein and lipid in the recombination mixture. In the electron microscope, the lipoprotein aggregates showed the typical trilaminar structure of membranes. Most of the aggregates formed closed vesicles. The same appearance was exhibited by the protein-free lipids used for the recombination experiment