Direct Electrochemistry of Shewanella oneidensis Cytochrome c Nitrite Reductase

Abstract

Shewanella oneidensis cytochrome c nitrite reductase (soNrfA) is a dimeric enzyme that plays a key role in bacterial nitrate respiration. The periplasmic reductase catalyzes the six‐electron reduction of nitrite to ammonia but can also catalyze the two‐electron reduction of hydroxylamine, a proposed intermediate in the nitrite reduction pathway. Here we use Protein Film Voltammetry to investigate the electrochemical and catalytic properties of soNrfA and compare its properties to the related Escherichia coli nitrite reductase (ecNrfA). Our findings highlight the similarities and differences between the two enzymes: most notably, all detected electron transfer steps are one‐electron in nature, contrary to what has been observed in ecNrfA. This suggests that for soNrfA the six‐electron reduction of nitrite is carried out in a series of one‐electron steps. Additionally we observe substrate inhibition during nitrite turnover and evidence of negative cooperativity during hydroxylamine turnover which suggests individual monomers within a ccNiR homodimer interact with one another during catalysis.