Characterization of unusual multicopper nitrite reductase containing blue and green type 1 Cu ions

Dissimilatory nitrite reductase isolated from Hyphomicrobium denitrificans A3151 (HdNIR) is a novel copper-containing nitrite reductase (CuNIR) composed of six identical subunits. One plastocyanin-like domain and one green CuNIR-like domain are connected to each other, suggesting that the HdNIR subunit structure resembles a complex of green CuNIR and pseudoazurin (or azurin). Recombinant HdNIR protein was crystallized using the hanging-drop vapour-diffusion method with PEG 4000 as the precipitant at pH 8.9. X-ray diffraction data were collected to 2.35 A resolution. The HdNIR crystal belonged to the tetragonal space group P4(1) (or P4(3)), with unit-cell parameters a = b = 221.9, c = 165.2 A, giving 12 molecules (two hexamers) per asymmetric unit and a solvent content of 64%. A mutant form of HdNIR, C260A, which lacks the type I copper ion in the CuNIR-like domain, was prepared and crystallized under wild-type HdNIR conditions. The C260A mutant crystal belonged to the cubic space group P4(3)32 (or P4(1)32), with unit-cell parameters a = b = c = 153.7 A, giving one molecule per asymmetric unit and a solvent content of 59%. X-ray diffraction data were collected to 3.5 A resolution. To solve the crystal structure of HdNIR, the multiwavelength anomalous dispersion (MAD) method and the molecular-replacement method are currently being used.

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Crystallization and preliminary X-ray crystallographic studies of dissimilatory nitrite reductase isolated fromHyphomicrobium denitrificansA3151

Xie

Inoue

Seike

et al. 2004

Acta Crystallogr D Biol Cryst

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Characterization of unusual multicopper nitrite reductase containing blue and green type 1 Cu ions

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Crystallization and preliminary X-ray crystallographic studies of dissimilatory nitrite reductase isolated fromHyphomicrobium denitrificansA3151

Crystallization and preliminary X-ray crystallographic studies of dissimilatory nitrite reductase isolated fromHyphomicrobium denitrificansA3151

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