TY - JOUR
T1 - Tetanus toxin interaction with human erythrocytes. II. Kinetic properties of toxin association and evidence for a ganglioside-toxin macromolecular complex formation
AU - Lazarovici, Philip
AU - Yavin, Ephraim
PY - 1985/1/25
Y1 - 1985/1/25
N2 - The properties of tetanus toxin interaction with human erythrocytes supplemented with disialo- and trisialo-gangliosides have been investigated. Binding of toxin is linear with time for 1 h and is 3-4-fold higher at 37°C than at 4°C during incubation of long duration. It exhibits saturation at toxin concentrations between 0.1 and 1 μg/ml; however, it is nonsaturable between 1 and up to 50 μg/ml. It is effectively prevented by free gangliosides and antibodies or by pretreatment with sialidase but is unaffected by a number of closely related ligands including toxoid and toxin fragments. NaCl (1 M) removes a great portion (86%) of cell-associated toxin while Triton X-100 extracts an additional fraction (30%) of the salt-resistant cell-bound toxin. The residual sequestred toxin after detergent extraction is sensitive to proteolytic degradation. The trypsin-stable fraction (1.5%) is biotoxic and may be indicative of internalization of toxin. A macromolecular complex of about 700 kDa containing toxin and gangliosides has been isolated characterized by Sephacryl S-300 gel permeation chromatography, SDS-gel electrophoresis, immunoprecipitability and biotoxicity. This complex is obtained only in ganglioside-supplemented cells and not when free 3H-labeled GDib is reacted with 125I-labeled toxin in solution in the absence of cells. The hydrophobicity properties acquired as a result of ganglioside-toxin interaction, presumably at the cell surface, suggest a conformational change of the toxin which may enable its penetration into the bilayer.
AB - The properties of tetanus toxin interaction with human erythrocytes supplemented with disialo- and trisialo-gangliosides have been investigated. Binding of toxin is linear with time for 1 h and is 3-4-fold higher at 37°C than at 4°C during incubation of long duration. It exhibits saturation at toxin concentrations between 0.1 and 1 μg/ml; however, it is nonsaturable between 1 and up to 50 μg/ml. It is effectively prevented by free gangliosides and antibodies or by pretreatment with sialidase but is unaffected by a number of closely related ligands including toxoid and toxin fragments. NaCl (1 M) removes a great portion (86%) of cell-associated toxin while Triton X-100 extracts an additional fraction (30%) of the salt-resistant cell-bound toxin. The residual sequestred toxin after detergent extraction is sensitive to proteolytic degradation. The trypsin-stable fraction (1.5%) is biotoxic and may be indicative of internalization of toxin. A macromolecular complex of about 700 kDa containing toxin and gangliosides has been isolated characterized by Sephacryl S-300 gel permeation chromatography, SDS-gel electrophoresis, immunoprecipitability and biotoxicity. This complex is obtained only in ganglioside-supplemented cells and not when free 3H-labeled GDib is reacted with 125I-labeled toxin in solution in the absence of cells. The hydrophobicity properties acquired as a result of ganglioside-toxin interaction, presumably at the cell surface, suggest a conformational change of the toxin which may enable its penetration into the bilayer.
KW - (Erythrocyte membrane)
KW - Cell surface
KW - Ganglioside
KW - Hydrophobic chromatography
KW - Membrane-toxin interaction
KW - Tetanus toxin
UR - http://www.scopus.com/inward/record.url?scp=0021978808&partnerID=8YFLogxK
U2 - 10.1016/0005-2736(85)90328-1
DO - 10.1016/0005-2736(85)90328-1
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C2 - 3967025
AN - SCOPUS:0021978808
SN - 0005-2736
VL - 812
SP - 532
EP - 542
JO - Biochimica et Biophysica Acta - Biomembranes
JF - Biochimica et Biophysica Acta - Biomembranes
IS - 2
ER -