Comparison of the structural characteristics of cell-CAM 105 from hepatocytes with those of an altered form expressed by rat transplantable hepatocellular carcinomas

The structural features of the adult rat hepatocyte (ARH) forms of cell-CAM 105, a Mr 105,000 cell adhesion molecule, were compared using a variety of immunochemical and biochemical techniques with altered forms of more basic pI present on two transplantable hepatocellular carcinomas (THC 1682c and THC AS-30D). Immunoprecipitation analysis with polyclonal (anti-gp 105-2) and monoclonal (MAb) antibodies specific for cell-CAM 105 (MAb 362.50) demonstrated that ARH and THC cell-CAM 105 were indistinguishable in several respects including: (a) binding to wheat germ agglutinin; (b) labeling with NaIO4/NaB3H4; (c) susceptibility to digestion with endoglycosidases (endoglycosidase H and F and peptide N-glycosidase F N-glycanase); (d) rate of turnover on the cell surface; and (e) differential resistance of upper and lower forms to trypsin digestion in the presence or absence of calcium. Digestion with Clostridium perfringens or Vibrio cholerae neuraminidase did not equalize pI but instead decreased the size and increased the pI of both ARH and THC cell-CAM 105. Comparison of two-dimensional tryptic peptide maps, however, revealed five unique peptides in the THC AS-30D map and one peptide in the THC 1682c map, peptides which were only apparent in maps of deglycosylated ARH cell-CAM 105. Based on these results, it was concluded that there were significant differences in the glycosylation of ARH and THC cell-CAM 105. Biosynthetic labeling with 32PO4 and 35SO4 showed that both ARH and THC molecules were phosphorylated but not sulfated. Comparison of 32P-labeled peptides produced by digestion with V-8 protease revealed significant differences in the phosphorylation of the upper and lower forms from ARH and showed that the pattern of phosphorylation on THC cell-CAM 105 most closely resembled ARH upper form. Pulse-chase analysis of ARH cell-CAM 105 further indicated that only a subpopulation of the molecules labeled with [35S]methionine were phosphorylated.