Occurrence of alpha-D-galactosyl-containing glycoproteins on Ehrlich tumor cell membranes

Ehrlich ascites tumor cells were strongly agglutinated by 0.4 micrograms/mL Griffonia simplicifolia I-B4 isolectin (GS I-B4), indicating the presence of nonreducing terminal alpha-D-galactopyranose (alpha-D-Galp) residues on the cell surface. Incubation of the cells with GS I-B4 labeled with either fluorescein isothiocyanate (FITC-B4) or ferritin followed by examination with the light and electron microscope revealed a random distribution of alpha-D-Galp residues over the entire cell surface. Cell-binding studies with [3H]propionate-labeled GS I-B4 demonstrated a minimum of 18.1 X 10(6) alpha-D-Galp sites per Ehrlich cell. An enriched Ehrlich cell plasma membrane preparation subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with FITC-B4 revealed a number of alpha-D-galactosyl-containing glycoproteins ranging in molecular weight from 50 000 to over 200 000. The major plasma membrane glycoprotein of the Ehrlich cell (GP 130) was isolated in near homogeneous form by using nonionic detergent extraction, affinity chromatography over GS I-Sepharose 4B, and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Injection of Ehrlich cells into the mouse peritoneal cavity stimulated the appearance of low levels of alpha-D-galactosyl-containing glycoproteins in the ascites fluid ranging in molecular weight from 34 000 to 260 000. These glycoproteins differed in molecular weight when compared to the alpha-D-galactosyl-containing glycoproteins observed in either ascites fluid induced with Freund's complete adjuvant or the glycoproteins in the Ehrlich cell plasma membrane.