Monoclonal antibody studies of ferredoxin:NADP+ oxidoreductase.
Eleven independent monoclonal antibodies, all IgG's, have been raised against the ferredoxin:NADP+ oxidoreductase of spinach leaves. All 11 monoclonal antibodies were able to produce substantial inhibition of the NADPH to 2,6-dichlorophenol indophenol (DCPIP) diaphorase activity of the enzyme, but none of the antibodies produced any significant inhibition of electron flow from NADPH to ferredoxin catalyzed by the enzyme. Spectral perturbation assays were used to demonstrate that antibody interaction with NADP+ reductase did not interfere significantly with the binding of either ferredoxin or NADP+ to the enzyme. Ultrafiltration binding assays were used to confirm that the monoclonal antibodies did not interfere with complex formation between ferredoxin and the enzyme. These results have been interpreted in terms of the likely presence of one or more highly antigenic epitopes at the site where the nonphysiological electron acceptor, DCPIP, binds to the enzyme. Furthermore, the results suggest that the site where DCPIP is reduced differs from both of the two separate sites at which the two physiological substrates, ferredoxin and NADP+/NADPH, are bound.
Chang
K T
KT
Department of Chemistry and Biochemistry, Texas Tech University, Lubbock 79409-1061.
Morrow
K J
KJ
Hirasawa
M
M
Knaff
D B
DB
D000911:Antibodies, Monoclonal; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D003574:Cytochrome c Group / Q000378:metabolism; D008058:Dihydrolipoamide Dehydrogenase / Q000378:metabolism; D004579:Electron Transport; D005287:Ferredoxin-NADP Reductase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000276:immunology; D009249:NADP / Q000378:metabolism; D010940:Plant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000276:immunology
D000911:Antibodies, Monoclonal; D003574:Cytochrome c Group; D010940:Plant Proteins; D009249:NADP; D005287:Ferredoxin-NADP Reductase; D008058:Dihydrolipoamide Dehydrogenase