Arginase and nitric oxide synthase (NOS) were inhibited by nitrite and putrescine, respectively. Results showed a cross-inhibition by the products of the two arginine utilizing pathways in macrophages. The kinetics of these inhibitions have been described earlier. Arginase was measured by the release of urea; NOS activity was determined by measuring 14C-L-citrulline synthesis from 14C-labeled L-arginine. The structural changes of arginase were studied by fluorescence and gel filtration experiments. Nitrite caused a decrease of tryptophane fluorescence over 5 mM concentration without dissociating the arginase oligomers as indicated by gel filtration experiments. The differences in the structural features of L-arginine substrate and putrescine inhibitor made the binding of putrescine to the active site of NOS unlikely. In conclusion, our studies suggest that nitrite causes a non-competitive inhibition of arginase based on a conformational change without the dissociation of the arginase oligomers. For putrescine inhibition we suggest an allosteric mechanism also based on a conformational change.

Keywords: macrophage, Arginase, NO synthase, Fluorescence, Gel Filtration, Conformational change