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A three-domain copper-nitrite reductase with a unique sensing loop.

ABSTRACT: Dissimilatory nitrite reductases are key enzymes in the denitrification pathway, reducing nitrite and leading to the production of gaseous products (NO, N₂O and N₂). The reaction is catalysed either by a Cu-containing nitrite reductase (NirK) or by a cytochrome cd ₁ nitrite reductase (NirS), as the simultaneous presence of the two enzymes has never been detected in the same microorganism. The thermophilic bacterium Thermus scotoductus 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 TsNirK has been determined at 1.63 Å resolution. TsNirK 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_N) while the C-terminus contains an extra type 1 Cu (T1Cu_C) bound within a cupredoxin motif. T1Cu_N shows an unusual Cu atom coordination (His₂-Cys-Gln) compared with T1Cu observed in NirKs reported so far (His₂-Cys-Met). T1Cu_C is buried at ∼5 Å from the molecular surface and located ∼14.1 Å away from T1Cu_N; T1Cu_N and T2Cu are ∼12.6 Å apart. All these distances are compatible with an electron-transfer process T1Cu_C → T1Cu_N → T2Cu. T1Cu_N and T2Cu are connected by a typical Cys-His bridge and an unexpected sensing loop which harbours a Ser_CAT residue close to T2Cu, suggesting an alternative nitrite-reduction mechanism in these enzymes. Biophysicochemical and functional features of TsNirK are discussed on the basis of X-ray crystallography, electron paramagnetic resonance, resonance Raman and kinetic experiments.

SUBMITTER: Opperman DJ

PROVIDER: S-EPMC6400189 | biostudies-literature |

REPOSITORIES: biostudies-literature

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