Interaction of concanavalin A with differentiated and undifferentiated murine neuroblastoma cells

Abstract

Tritium-labeled acetyl-concanavalin A ( 3H-Con A) was used to study its kinetics of binding at 0 °C to murine neuroblastoma cells (clone neuro 2-A) grown in the differentiated (monolayer) and Undifferentiated (spinner) states. The binding of 3H-Con A to both cell types gives sigmoidal saturation curves, suggesting positively cooperative binding of the lectin. The Hill coefficient is 1.75 for differentiated and 1.36 for Undifferentiated cells. The maximal number of 3H-Con A molecules bound per cell is 2.3 × 10 7 and 3.4 × 10 7 for differentiated and Undifferentiated cells, respectively, and the apparent rate constants for formation of the lectin-cell complex are 6.13 × 10 2, m −1, s −1 for the Undifferentiated and 6.68 × 10 2, m −1, s −1 for the differentiated cells. The lectin bound to spinner cells does not dissociate spontaneously to any measurable extent over a 60-min period at 0 or 37 °C, but the lectin-cell complex dissociates rapidly after addition of α-methyl- d-mannopyranoside. At 37 °C, this sugar causes virtually complete dissociation of the cell-lectin complex within 30 min. The 3H-Con A dissociated from spinner cells is indistinguishable from the original 3H-Con A by sodium dodecyl sulfate-urea polyacrylamide gel electrophoresis, gel filtration through Bio-Gels P-30 and P-100, and specific binding to spinner cells. Both the original and the dissociated 3H-Con A are dimers at pH 7.4. The sugar-induced dissociation of the labeled lectin from spinner cells is not accompanied by shedding or inactivation of the lectin binding sites of the cell surface.