Denitrification by the fungus Fusarium oxysporum and involvement of cytochrome P-450 in the respiratory nitrite reduction

From conditions for production in Fusarium oxysporum of the unique nitrate/nitrite-inducible cytochrome P-450, tentatively called P-450dNIR, it was expected that the fungus is capable of metabolizing nitrate dissimilatively. Here we report that F. oxysporum exhibits a distinct denitrifying ability which results in the anaerobic evolution of nitrous oxide (N2O) from nitrate or nitrite. Comparison of the cell growth during denitrification indicated that the dissimilatory reduction of nitrate to nitrite is an energetically favorable process in F. oxysporum; however, further reduction of nitrite to N2O might be energy-exhausting and may function as a detoxification mechanism. A potent nitrite reductase activity to form N2O could be reconstituted by combination of the cell-free extract prepared from the denitrifying cells and an NADH-phenadinemethosulfate-dependent reducing system. The activity was strongly inhibited by carbon monoxide, cyanide, oxygen (O2), and the antibody against P-450dNIR. The results, along with those concerning inducing conditions of P-450dNIR, were highly indicative that the cytochrome is involved in the denitrifying nitrite reduction. This work has thus presented not only the first demonstration that a eukaryote exhibits a marked denitrifying ability, but also the first instance of a cytochrome P-450 that is involved in a reducing reaction with a distinct physiological significance against a hydrophilic, inorganic substrate.