Monoclonal antibody probes for the niaD specified subunit in the NADPH-nitrate reductase from Aspergillus nidulans.
In Aspergillus nidulans, the nitrate assimilatory pathway is regulated by a variety of agents, one being the autogenous enzyme nitrate reductase. A major subunit of the enzyme which is specified by the niaD structural gene and is implicated in autogenous control exhibits both nitrate inducible diaphorase activity and ammonium repression. The former was used to test the extent to which alterations in the niaD specified protomer might affect its formation in selected niaD point and deletion mutants. Enzyme preparations from the wild type and mutant strains were compared on the basis of nitrate inducible co-activities and their reaction to specific monoclonal antibodies (MABS). Proteins in partially purified mycelial extracts were subjected to Western blot analyses with three MABs to functional native enzyme. Extracts of niaD point mutants exhibited nitrate induced co-activities which matched those of the wild type while those from deletion mutants were diminished or totally inactive. Nitrate reductase, from the wild type and specific cofactor mutants, shares an epitope common to both the monomeric and dimeric form in the case of one MAB, and exhibits epitopes unique to one or the other form in the case of the other two forms. Enzyme-antibody interaction occurs with or without inhibition of catalytic activity depending on the MAB involved.
Downey
R J
RJ
Department of Microbiology, Ohio University, Athens 45701.
Cook
J A
JA
Brumfield
A M
AM
D000818:Animals; D000911:Antibodies, Monoclonal; D001233:Aspergillus nidulans / Q000201:enzymology* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005796:Genes; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D050896:Nitrate Reductase (NAD(P)H); D009565:Nitrate Reductases / Q000235:genetics / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D009565:Nitrate Reductases; D050896:Nitrate Reductase (NAD(P)H)