title: Characterization of tetanus toxin binding to rat brain membranes. Evidence for a high-affinity proteinase-sensitive receptor.
abstract: Binding of 125I-labelled tetanus toxin to rat brain membranes in 25 mM-Tris/acetate, pH 6.0, was saturable and there was a single class of high-affinity site (KD 0.26-1.14 nM) present in high abundance (Bmax. 0.9-1.89 nmol/mg). The sites were largely resistant to proteolysis and heating but were markedly sensitive to neuraminidase. Trisialogangliosides were effective inhibitors of toxin binding (IC50 10 nM) and trisialogangliosides inserted into membranes lacking a toxin receptor were able to bind toxin with high affinity (KD 2.6 nM). The results are consistent with previous studies and the hypothesis that di- and trisialogangliosides act as the primary receptor for tetanus toxin under these conditions. In contrast, when toxin binding was assayed in Krebs-Ringer buffer, pH 7.4, binding was greatly reduced, was non-saturable and competition binding studies showed evidence for a small number of high-affinity sites (KD 0.42 nM, Bmax. 0.90 pmol/mg) and a larger number of low-affinity sites (KD 146 nM, Bmax. 179 pmol/mg). Treatment of membranes with proteinases, heat, and neuraminidase markedly reduced binding. Trisialogangliosides were poor inhibitors of toxin binding (IC50 11.0 microM), and trisialogangliosides inserted into membranes bound toxin with low affinity. The results suggest that in physiological buffers tetanus toxin binds with high affinity to a protein receptor, and that gangliosides represent only a low-affinity site.
mesh_terms: D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000378:metabolism; D006224:Cricetinae; D005732:Gangliosides / Q000494:pharmacology; D007668:Kidney / Q000378:metabolism; D008565:Membrane Proteins; D010447:Peptide Hydrolases; D051381:Rats; D011919:Rats, Inbred Strains; D011950:Receptors, Cholinergic / Q000187:drug effects / Q000378:metabolism*; D013744:Tetanus Toxin / Q000378:metabolism*
chemical_list: D005732:Gangliosides; D008565:Membrane Proteins; D011950:Receptors, Cholinergic; D013744:Tetanus Toxin; C028728:tetanus toxin receptor; C016806:trisialoganglioside; D002772:Cholera Toxin; D010447:Peptide Hydrolases