Partial characterization of fucosylated cell surface glycoproteins of cultured RPE.
The major high molecular weight, fucose containing, cell surface glycoproteins of cultured rat retinal pigment epithelial (RPE) cells were partially characterized. One dimensional peptide mapping by the Cleveland method showed that the polypeptide chains of these proteins were not highly related in structure. Incorporation of 3H-mannose into these glycoproteins was equivalent for normal and dystrophic (RCS rdy-p+) RPE. Furthermore, treatment of the glycoproteins from either normal or dystrophic RPE with Endo-beta-N-acetylglucosaminidase H (Endo H) did not cause a shift in their Mr's, as determined by SDS PAGE. These results suggest that the high Mr glycoproteins do not contain a large quantity of unprocessed, mannose containing core type N-linked oligosaccharides in either normal or dystrophic RPE. Digestion of the 3H-fucose labeled glycoproteins with Peptide N-glycosidase F (PNGase F) demonstrated that at least 90% of the 3H-fucose incorporated into these glycoproteins is in N-linked oligosaccharides. Endo-beta-N-acetylglucosaminidase F (Endo F) treatment showed that at least 75-80% of the 3H-fucose is located in more terminal positions (distal to the fucose that is found in alpha 1,6 linkage to the asparagine-linked N-acetylglucosamine residue) in N-linked carbohydrate. Overall, these results support the hypothesis that if the dystrophic RPE possesses a defect in glycoprotein processing, then this defect affects terminal processing of oligosaccharides and addition of terminally located fucose residues. A homologous group of high Mr, fucosylated glycoproteins was found in plasma membranes from cultured monkey RPE, suggesting atht they may be common to other species.
Clark
V M
VM
Jules Stein Eye Institute, Los Angeles, CA 90024-7008.
Zhou
X Y
XY
Pfeffer
B A
BA
D000118:Acetylglucosaminidase; D000581:Amidohydrolases; D000818:Animals; D002240:Carbohydrate Sequence; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D005643:Fucose / Q000378:metabolism*; D006596:Hexosaminidases; D008253:Macaca mulatta; D008358:Mannose / Q000378:metabolism; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008562:Membrane Glycoproteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010449:Peptide Mapping; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D010857:Pigment Epithelium of Eye / Q000737:chemistry*; D051381:Rats; D011919:Rats, Inbred Strains; D011922:Rats, Mutant Strains
D008562:Membrane Glycoproteins; D005643:Fucose; D006596:Hexosaminidases; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D008358:Mannose