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PDBsum entry 2nya
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Oxidoreductase
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PDB id
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2nya
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References listed in PDB file
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Key reference
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Title
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Spectropotentiometric and structural analysis of the periplasmic nitrate reductase from escherichia coli.
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Authors
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B.J.Jepson,
S.Mohan,
T.A.Clarke,
A.J.Gates,
J.A.Cole,
C.S.Butler,
J.N.Butt,
A.M.Hemmings,
D.J.Richardson.
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Ref.
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J Biol Chem, 2007,
282,
6425-6437.
[DOI no: ]
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PubMed id
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Abstract
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The Escherichia coli NapA (periplasmic nitrate reductase) contains a [4Fe-4S]
cluster and a Mo-bis-molybdopterin guanine dinucleotide cofactor. The NapA
holoenzyme associates with a di-heme c-type cytochrome redox partner (NapB).
These proteins have been purified and studied by spectropotentiometry, and the
structure of NapA has been determined. In contrast to the well characterized
heterodimeric NapAB systems ofalpha-proteobacteria, such as Rhodobacter
sphaeroides and Paracoccus pantotrophus, the gamma-proteobacterial E. coli NapA
and NapB proteins purify independently and not as a tight heterodimeric complex.
This relatively weak interaction is reflected in dissociation constants of 15
and 32 mum determined for oxidized and reduced NapAB complexes, respectively.
The surface electrostatic potential of E. coli NapA in the apparent NapB binding
region is markedly less polar and anionic than that of the alpha-proteobacterial
NapA, which may underlie the weaker binding of NapB. The molybdenum ion
coordination sphere of E. coli NapA includes two molybdopterin guanine
dinucleotide dithiolenes, a protein-derived cysteinyl ligand and an oxygen atom.
The Mo-O bond length is 2.6 A, which is indicative of a water ligand. The
potential range over which the Mo(6+) state is reduced to the Mo(5+) state in
either NapA (between +100 and -100 mV) or the NapAB complex (-150 to -350 mV) is
much lower than that reported for R. sphaeroides NapA (midpoint potential
Mo(6+/5+) > +350 mV), and the form of the Mo(5+) EPR signal is quite distinct.
In E. coli NapA or NapAB, the Mo(5+) state could not be further reduced to
Mo(4+). We then propose a catalytic cycle for E. coli NapA in which nitrate
binds to the Mo(5+) ion and where a stable des-oxo Mo(6+) species may
participate.
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Figure 5.
FIGURE 5. Crystal structure of E. coli NapA. A, view of
NapA from E. coli with the different domains colored as follows:
domain I (residues 1–59, 492–520, and 590–630) in red;
domain II (residues 60–138, 375–491, and 521–589) in
green; domain III (residues 139–374) in yellow; and domain IV
(residues 631–791) in blue. The regions colored gray are the
extra loop regions not present in the D. desulfuricans NAP (9).
The [4Fe-4S] cluster and the Mo-bis-MGD cofactors are shown in
ball and stick format and are colored black. B, view of the
4Fe4S cluster and Mo-bis-MGD cofactor showing the conserved
water molecule that is coordinated by Lys-47 and one of the MGD
moieties.
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Figure 6.
FIGURE 6. Electrostatic surface representations of the NAP
structures. A, electrostatic surface of the molybdenum funnel
face of the various NAP structures. B, electrostatic surface of
the presumed NapB interaction face of the various NAP
structures. Electrostatic surfaces were calculated in the
program GRASP (28) and displayed in PYMOL (Delano Scientific).
The "surface potentials" of color property scale -10 (red,
negatively charged) to +10 (blue, positively charged).
Coordinates for D. desulfuricans NAP, PDB file 2NAP. Coordinates
for R. sphaeroides NapA, Protein Data Bank (PDB) file 1OGY. The
structures of E. coli, D. desulfuricans, and R. sphaeroides were
aligned using Swisspdbviewer 3.0 ("Magic Fit"). D.d, D.
desulfuricans; E.c, E. coli, R.s, R. sphaeroides.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2007,
282,
6425-6437)
copyright 2007.
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