Mapping of the monoclonal antibody 80L5C4 epitope on the cell adhesion molecule gp80 of Dictyostelium discoideum.
EDTA-resistant cell-cell binding sites are expressed on Dictyostelium discoideum cells at the aggregation stage of development. A cell surface glycoprotein of Mr 80,000 (gp80) has been found to mediate these binding sites via homophilic interaction. We have previously raised a monospecific monoclonal antibody 80L5C4 against gp80, which blocks the cell binding site of gp80 (Siu, C.-H., Lam, T.Y. and Choi, A.H.C. (1985) J. Biol. Chem. 260, 16030-16036). To map the 80L5C4 epitope, gp80 was digested with protease V8, and the smallest proteolytic fragment that retained immunoreactivity with 80L5C4 was about 27,000 Da, corresponding to the amino-terminal fragment predicted from the cleavage sites. In addition, cDNA fragments containing different gp80 coding regions were used to construct trpE/gp80 gene fusions in the expression vector pATH10. An analysis of these fusion proteins led to the mapping of the 80L5C4 epitope to a 51 amino-acid segment between residues 123 and 173.
Kamboj
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Banting and Best Department of Medical Research, University of Toronto, Canada.
Siu
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CH
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000954:Antigens, Surface / Q000276:immunology*; D001665:Binding Sites; D002448:Cell Adhesion / Q000187:drug effects; D015815:Cell Adhesion Molecules; D004023:Dictyostelium / Q000032:analysis*; D004492:Edetic Acid / Q000494:pharmacology; D000939:Epitopes / Q000032:analysis* / Q000276:immunology; D004926:Escherichia coli / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000276:immunology; D012697:Serine Endopeptidases / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D000939:Epitopes; D010446:Peptide Fragments; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C052291:cell cohesion molecule, Dictyostelium; D004492:Edetic Acid; D010447:Peptide Hydrolases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase