Dissimilatory nitrite reductases are key enzymes in the denitrification pathway， reducing nitrite and leading to the production of gaseous products (NO， NO and N). The reaction is catalysed either by a Cu-containing nitrite reductase (NirK) or by a cytochrome nitrite reductase (NirS)， as the simultaneous presence of the two enzymes has never been detected in the same microorganism. The thermophilic bacterium SA-01 is an exception to this rule， harbouring both genes within a denitrification cluster， which encodes for an atypical NirK. The crystal structure of NirK has been determined at 1.63 Å resolution. NirK is a homotrimer with subunits of 451 residues that contain three copper atoms each. The ... More
Dissimilatory nitrite reductases are key enzymes in the denitrification pathway， reducing nitrite and leading to the production of gaseous products (NO， NO and N). The reaction is catalysed either by a Cu-containing nitrite reductase (NirK) or by a cytochrome nitrite reductase (NirS)， as the simultaneous presence of the two enzymes has never been detected in the same microorganism. The thermophilic bacterium SA-01 is an exception to this rule， harbouring both genes within a denitrification cluster， which encodes for an atypical NirK. The crystal structure of NirK has been determined at 1.63 Å resolution. NirK is a homotrimer with subunits of 451 residues that contain three copper atoms each. The N-terminal region possesses a type 2 Cu (T2Cu) and a type 1 Cu (T1Cu) while the C-terminus contains an extra type 1 Cu (T1Cu) bound within a cupredoxin motif. T1Cu shows an unusual Cu atom coordination (His-Cys-Gln) compared with T1Cu observed in NirKs reported so far (His-Cys-Met). T1Cu is buried at ∼5 Å from the molecular surface and located ∼14.1 Å away from T1Cu; T1Cu and T2Cu are ∼12.6 Å apart. All these distances are compatible with an electron-transfer process T1Cu → T1Cu → T2Cu. T1Cu and T2Cu are connected by a typical Cys-His bridge and an unexpected sensing loop which harbours a Ser residue close to T2Cu， suggesting an alternative nitrite-reduction mechanism in these enzymes. Biophysicochemical and functional features of NirK are discussed on the basis of X-ray crystallography， electron paramagnetic resonance， resonance Raman and kinetic experiments.
