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Figure 4
Proposed Cu–CAII nitrite reductase mechanism. (a) Cu–CAII in the resting state with a copper-bound solvent molecule. Thr199 is slightly acidic because of interactions with Glu106 allowing Thr199 to stabilize the solvent molecule. W1 is stabilized via hydrogen bonding to Thr200 and W2 (not shown for clarity). (b) NO2− enters the active site, displacing the copper-bound solvent. NO2− binds in a `hat' conformation, with both oxygen atoms coordinating to the copper. One oxygen is primed for catalysis via hydrogen bonding to the hydroxyl of Thr199 and W1. (c) Intermolecular electron transfer from the T-1 copper site, donating an electron to the T-2 copper site and generating a Cu+ cation in the T-2 active site. This triggers a binding-mode change in NO2− from `hat' to `side-on' coordination. One oxygen is uncoordinated from the copper and stabilized via the nitro­gen from Thr199 while the other oxygen retains hydrogen bonding to Thr199 and W1. (d) Reduction of nitrite begins via an electron donation from Cu+, resulting in a cascade of electron rearrangement and the regeneration of Cu2+. The primed oxygen accepts two protons from W1 and the acidic hydroxyl of Thr199 forming a bound water molecule. (e) The nitrite molecule is now reduced to nitric oxide and transiently bound to the Cu2+ cation along with the generated water molecule. As the water molecule forms, the nitric oxide is released from the copper. More protons are shuttled into the active site via the CA proton-shuttle His64 and the necessary catalytic protons are replenished regenerating the resting state in (a) (Duda et al., 2001BB13).

IUCrJ
Volume 7| Part 2| March 2020| Pages 287-293
ISSN: 2052-2525
https://doi.org/10.1107/S2052252520000986