Development of monoclonal antibodies against different protein and carbohydrate epitopes of dipeptidyl peptidase IV from rat liver plasma membranes.
Dipeptidyl peptidase IV (DPP IV) is a serine exopeptidase expressed at high levels in rat kidney, liver and lung. We established eight monoclonal antibodies against partially purified DPP IV from rat liver plasma membranes. By means of a competitive dot blot assay with purified DPP IV, these antibodies were shown to recognize four different epitopes of the glycoprotein, designated A - D. The epitopes are located on the extracellular domain of DPP IV, as shown by papain digestion of liver plasma membranes. Treatment of DPP IV with neuraminidase and glycopeptide N-glycosidase F, as well as incubation of hepatocytes with the alpha-mannosidase I inhibitor deoxymannojirimycin, revealed that epitope A may be formed by a mannose-rich sugar chain and epitope D might represent a complex carbohydrate structure in the mature glycoprotein, while the epitopes B and C are formed by the protein moiety. Concanavalin A reduced the binding of monoclonal antibody to epitope A by 78%. Binding to epitope D was blocked by 73% with wheat germ lectin, and by more than 99% with sialic acid; epitopes B and C were unaffected by any of the lectins or sugars tested. The immunological cross-reactivity with DPP IV from Morris hepatoma 7777 was demonstrated with monoclonal antibodies against epitopes A-C. Epitope D was not recognized on hepatoma DPP IV. However, in addition to DPP IV, four hepatoma plasma membrane glycoproteins were precipitated by the monoclonal antibody against the epitope D, indicating that this epitope is not uniquely restricted to DPP IV.
Hartel-Schenk
S
S
Institut für Molekularbiologie und Biochemie, Freie Universität Berlin, Federal Republic of Germany.
Loch
N
N
Zimmermann
M
M
Reutter
W
W
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D001667:Binding, Competitive / Q000276:immunology; D002241:Carbohydrates / Q000276:immunology*; D002462:Cell Membrane / Q000201:enzymology*; D003429:Cross Reactions; D018819:Dipeptidyl Peptidase 4; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000276:immunology* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D006023:Glycoproteins / Q000276:immunology; D006031:Glycosylation; D037102:Lectins / Q000378:metabolism; D008099:Liver / Q000201:enzymology*; D008114:Liver Neoplasms, Experimental / Q000201:enzymology; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D002241:Carbohydrates; D000939:Epitopes; D006023:Glycoproteins; D037102:Lectins; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D018819:Dipeptidyl Peptidase 4