Contributions of lipids and proteins to the surface charge of membranes. An electron microscopy study with cationized and anionized ferritin.

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Abstract
The surface charge of cultured neurons was investigated with EM markers anionized ferritin (AF) and cationized ferritin (CF). To determine which membrane components react with the markers, model reactions were used. Protein-coated Sepharose beads and lipid vesicles were reacted at physiological pH. Groups which may contribute to the surface charge are acidic groups, the sialic acid of both glycoproteins and gangliosides, the carboxyl group of proteins and the phosphates of phospholipids; basic groups, the amines of proteins. The effect of chemical fixation on surface charge was investigated. Glutaraldehyde fixation increased the charge of neutral proteins but not by a mechanism involving unbound aldehydes. Glutaraldehyde fixation of phospholipid vesicles in the presence of CF showed that amine-containing phospholipids were cross-linked to CF. This cross-linking was seen with EM as the clumping of CF and the burying of CF in the membrane. Paraformaldehyde fixation had a lesser effect on the charge of proteins but did react with phospholipids as did glutaraldehyde. At physiological pH, most of the charged proteins and lipids on cell surface contribute to the membrane surface charge and the membrane surface charge of cells can be greatly changed by chemical fixation.