Bacterial cytochrome c nitrite reductase: new structural and functional aspects

Cytochrome c nitrite reductase catalyzes the six-electron reduction of nitr ite to ammonia as a key step within the biological nitrogen cycle. Most rec ently, the crystal structure of the soluble enzyme from Sulfurospirillum de leyianum could be solved to 1.9 Angstrom resolution. This set the basis for new experiments on structural and functional aspects of the pentaheme prot ein which carries a Ca(2+) ion close to the active site heme. In the crysta l, the protein was a homodimer with ten hemes in very close packing. The st rong interaction between the nitrite reductase monomers also occurred in so lution according to the dependence of the activity on the protein concentra tion. Addition of Ca(2+) to the enzyme as isolated had a stimulating effect on the activity. Ca(2+) could be removed from the enzyme by treatment with chelating agents such as EGTA or EDTA which led to a decrease in activity. In addition to nitrite, the enzyme converted NO, hydroxylamine and O-methy l hydroxylamine to ammonia at considerable rates. With N2O the activity was much lower; most likely dinitrogen was the product in this case. Cytochrom e c nitrite reductase exhibited a remarkably high sulfite reductase activit y, with hydrogen sulfide as the product. A paramagnetic Fe(LT)-NO, S=1/2 ad duct was identified by rapid freeze EPR spectroscopy under turnover conditi ons with nitrite. This potential reaction intermediate of the reduction of nitrite to ammonia was also observed with PAPA NONOate and Spermine NONOate . (C) 2000 Elsevier Science Inc. All rights reserved.