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PDBsum entry 2ppf
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Oxidoreductase
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PDB id
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2ppf
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References listed in PDB file
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Key reference
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Title
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Stable copper-Nitrosyl formation by nitrite reductase in either oxidation state.
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Authors
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E.I.Tocheva,
F.I.Rosell,
A.G.Mauk,
M.E.Murphy.
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Ref.
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Biochemistry, 2007,
46,
12366-12374.
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PubMed id
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Abstract
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Nitrite reductase (NiR) is an enzyme that uses type 1 and type 2 copper sites to
reduce nitrite to nitric oxide during bacterial denitrification. A
copper-nitrosyl intermediate is a proposed, yet poorly characterized feature of
the NiR catalytic cycle. This intermediate is formally described as Cu(I)-NO+
and is proposed to be formed at the type 2 copper site after nitrite binding and
electron transfer from the type 1 copper site. In this study, copper-nitrosyl
complexes were formed by prolonged exposure of exogenous NO to crystals of
wild-type and two variant forms of NiR from Alcaligenes faecalis (AfNiR), and
the structures were determined to 1.8 A or better resolution. Exposing oxidized
wild-type crystals to NO results in the reverse reaction and formation of
nitrite that remains bound at the active site. In a type 1 copper site mutant
(H145A) that is incapable of electron transfer to the type 2 site, the reverse
reaction is not observed. Instead, in both oxidized and reduced H145A crystals,
NO is observed bound in a side-on manner to the type 2 copper. In AfNiR, Asp98
forms hydrogen bonds to both substrate and product bound to the type 2 Cu. In
the D98N variant, NO is bound side-on but is more disordered when observed for
the wild-type enzyme. The solution EPR spectra of the crystallographically
characterized NiR-NO complexes indicate the presence of an oxidized type 2
copper site and thus are interpreted as resulting from stable copper-nitrosyls
and formally assigned as Cu(II)-NO-. A reaction scheme in which a second NO
molecule is oxidized to nitrite can account for the formation of a Cu(II)-NO-
species after exposure of the oxidized H145A variant to NO gas.
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