Direct voltammetry of adsorbed redox enzymes at pyrolytic graphite electrodes has shown to be very useful to probe the catalytic activity of several nitrate reductases in the presence of nitrate. In this work we demonstrated that in some cases an electrode-induced haem alteration leads to a loss of nitrate reductase activity.Nitrate reductases are key enzymes in the biological nitrogen cycle. In particular, NapAB from Cupriavidus necator has an important role in the scavenging of nitrate and NarGHI from Marinobacter hydrocarbonoclasticus 617 is essential for the anaerobic respiration. These enzymes present haem groups among their redox centres, which are essential for the electron transfer from the reducing to the oxidising substrate. When adsorbed at graphite electrodes, both enzymes displayed a non-turnover signal corresponding to a one-electron redox process, with formal reduction potentials at pH 7.6 of -159 mV and -139 mV vs. SHE for Nap and Nar, respectively. Both enzymes displayed peroxidase activity at a potential close to that of the non-turnover response. Experiments with the whole enzymes and the haem free NapA from Desulfovibrio desulfuricans and NarGH from M. hydrocarbonoclasticus 617 were a valuable tool to get information about the cofactor undergoing electron transfer. We confirmed that this behaviour is related with the haems present in subunit B and subunit I of NapAB and NarGHI, respectively.
- Direct voltammetry
- Periplasmic nitrate reductase
- Peroxidase activity
- Respiratory nitrate reductase