Immunochemistry of Thioredoxin

Antisera against thioredoxin from Escherichia coli and phage T4 were prepared by immunization of rabbits. Both proteins were strongly immunogenic and produced specific precipitating antibodies with titers from 2 to 6 mg of antibody per ml of serum. The thioredoxin antibody of the immune rabbit serum is contained in the IgG-fraction. Antibodies against E. coli thioredoxin specifically inhibited the oxidation-reduction activity of the protein in the presence of thioredoxin reductase and NADPH. No cross-reactivity with T4 thioredoxin was observed in precipitin reactions or assays of oxidation-reduction activity. Cross-reaction with yeast thioredoxin was observed in oxidation-reduction assays and calculated to be around 25%. Yeast thioredoxin did not precipitate with antibodies against E. coli thioredoxin but inhibited the rate of precipitation of E. coli thioredoxin-antithioredoxin indicating identity of at least one single antigenic determinant in the two thioredoxins. Antibodies against T4 thioredoxin specifically inhibited T4 thioredoxin oxidation-reduction activity and showed no cross-reaction with E. coli or yeast thioredoxin in precipitin tests or enzymatic assays. The results showed the absence of common antigenic determinants in E. coli and T4 thioredoxin despite the fact that both proteins are substrates for the specific enzyme thioredoxin reductase of E. coli.