As a Fe^II-NO Mb complex is a possible intermediate, we compared the reduction of nitrite with NO(g). Significant differences are seen in the voltammograms. The Fe^III/IIwave is absent after the initial scan, due to the ready formation of Fe^II-NO, which is not oxidized under these conditions. A strong catalytic wave appears that is well positive to that of nitrite reduction. And in contrast to nitrite reduction, this first NO reduction is catalytic at all pH tested. Mass spectral analysis showed formation of N₂O at potentials associated with the first catalytic wave.

These results suggest the first wave in nitrite reduction includes both the reductive dehydration to form the nitrosyl, and its subsequent reduction to the nitroxyl (NO^-) adduct.

Catching the Nitroxyl

As we increase scan rate from 200 mV to 10 V/sec, we see that NO reductions are reversible at faster timescales-- the nitroxyl (NO^-) adduct is a distinguishable intermediate with a lifetime of ca. 100 milliseconds at pH 7. This reversibilitydecreases with increasing [H⁺] and [NO], which implies that thenitroxyl decomposes by reaction with these species.

We have electrochemical evidence for each step shown, but major questions remain:

--How does the protein active site influence the different steps? For example,consider the different heme structures in the native enzymes; will the siroheme nitrosyls be easier to reduce? Will they form a more stable nitoxyl adduct?

-- What differentiates between the two reductive pathways past nitroxyl? Does the three electron path occur through an initial dehydration-- i.e. a nitrido or nitrene intermediate?

To answer these and other questions, we are expanding our investigations using other hemeproteins....

Research | Home | Photochem | Electrochem | Presentations