title: Anionic sites of human erythrocyte membranes. II. Antispectrin-induced transmembrane aggregation of the binding sites for positively charged colloidal particles.
abstract: The effects of affinity-purified antispectrin gamma-globulins on the topographic distribution of anionic residues on human erythrocytes membranes was investigated using collo ida iron hydroxide labeling of mounted, fixed, ghost membranes. Antispectrin gamma-globulins were sequestered inside ghosts by hemolysis and the ghosts were incubated for 30 min at 37 degrees C and then fixed with glutaraldehyde. The topographic distribution of colloidal iron hydroxide clusters on ghosts incubated with low (<0.05 mg/ml) or high (>5-10 mg/ml concentrations of sequestered antispectrin was dispersed, but the distribution at intermediate concentrations (0.1-5 mg/ml) was highly aggregated. The aggregation of colloidal iron hydroxide binding sites was time and temperature dependent and required the sequestering of cross-linking antibodies (antispectrin Fab could not substitute for gamma-globulin antibodies) inside the ghosts. Prior glutaraldehyde fixation or fixation at the time of hemolysis in antispectrin solutions prevented the antispectrin-induced colloidal iron site aggregation. The antispectrin reacted exclusively at the inner ghost membrane surface and the colloidal iron hydroxide bound to N-acetylneuraminic acid residues on the outer membrane surface which are overwhelming on the sialoglycoprotein glycophorin. These results were interpreted as evidence for a structural transmembrane linkage between the inner surface peripheral protein spectrin and the integral membrane component glycophorin.
mesh_terms: D001665:Binding Sites; D001798:Blood Proteins / Q000378:metabolism; D002449:Cell Aggregation; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D004912:Erythrocytes / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007128:Immunoglobulin Fragments; D008854:Microscopy, Electron; D011485:Protein Binding; D011955:Receptors, Drug; D005719:gamma-Globulins / Q000378:metabolism
chemical_list: D001798:Blood Proteins; D007128:Immunoglobulin Fragments; D011955:Receptors, Drug; D005719:gamma-Globulins