Electrophoretic and Chemical Characterization of the Charged Groups at the Surface of Murine CL3 Ascites Leukaemia Cells

Abstract

The electrophoretic characteristics of the murine CL3 ascites tumour were investigated. Treatment of the cells with formaldehyde raised the electrophoretic mobility (E.P.M.) from - 1.06 to - 1.28 µ/sec/V/cm; subsequent treatment with diazomethane reduced their mobility to zero. The E.P.M. of the diazomethane-treated cells did not alter over the pH range 3.0-8.0. This proved that the only ionic groups at this cell surface were amino and carboxyl groups. The absence of phosphate groups, another possibility, was confirmed by the lack of calcium-ion binding from 10 mM Ca2+ solutions. Neuraminidase treatment reduced the E.P.M. from -1.06 to -0.55 µ/sec/V/cm and free sialic acid was identified in the enzyme supernatant. Subsequent treatment of the cells with formaldehyde raised the mobility to -1.22 µ/sec/V/cm indicating that the change in E.P.M. on neuraminidase treatment was not due solely to the removal of the carboxyl groups of sialic acid but also to a change in the ionic nature of the surface. This change is ascribed to a change in the conformation of the surface protein. The reason for this change and a suggestion for the possible role of sialic acid at the cell surface are mentioned. Treatment of the cells with trypsin did not affect the viable cells in any way, suggesting that the surface proteins lack the basic amino acids lysine and arginine. Pronase treatment served only to show that much of the sialic acid was bound to protein; the total amount was not determined.