 |
PDBsum entry 1df1
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Oxidoreductase
|
PDB id
|
|
|
|
1df1
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
Contents |
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
* Residue conservation analysis
|
|
|
|
|
References listed in PDB file
|
|
|
Key reference
|
|
|
Title
|
|
N-Terminal domain swapping and metal ion binding in nitric oxide synthase dimerization.
|
|
|
Authors
|
|
B.R.Crane,
R.J.Rosenfeld,
A.S.Arvai,
D.K.Ghosh,
S.Ghosh,
J.A.Tainer,
D.J.Stuehr,
E.D.Getzoff.
|
|
|
Ref.
|
|
EMBO J, 1999,
18,
6271-6281.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
Nitric oxide synthase oxygenase domains (NOS(ox)) must bind tetrahydrobiopterin
and dimerize to be active. New crystallographic structures of inducible NOS(ox)
reveal that conformational changes in a switch region (residues 103-111)
preceding a pterin-binding segment exchange N-terminal beta-hairpin hooks
between subunits of the dimer. N-terminal hooks interact primarily with their
own subunits in the 'unswapped' structure, and two switch region cysteines (104
and 109) from each subunit ligate a single zinc ion at the dimer interface.
N-terminal hooks rearrange from intra- to intersubunit interactions in the
'swapped structure', and Cys109 forms a self-symmetric disulfide bond across the
dimer interface. Subunit association and activity are adversely affected by
mutations in the N-terminal hook that disrupt interactions across the dimer
interface only in the swapped structure. Residue conservation and electrostatic
potential at the NOS(ox) molecular surface suggest likely interfaces outside the
switch region for electron transfer from the NOS reductase domain. The
correlation between three-dimensional domain swapping of the N-terminal hook and
metal ion release with disulfide formation may impact inducible nitric oxide
synthase (i)NOS stability and regulation in vivo.
|
|
|
|
|
|
|
|
Figure 1.
Figure 1 The effect of domain swapping on the N-terminal hook
conformation in iNOS[ox]. Ribbon representation of the iNOS[ox]
dimer in swapped (A) and unswapped (B) conformations. N-terminal
hook regions (cyan and orange) interact primarily with their own
subunits (purple and red) in the unswapped conformation, but
reach across to associate with the opposite subunit in the
swapped conformation. Each heme (yellow bonds) is cupped in the
inward-facing palm of the central webbed  -sheet
of the 'catcher's mitt' subunit fold. A self-symmetric disulfide
bond (yellow, bottom center) links the two subunits in the
swapped conformation (A). A single zinc ion (gray, bottom
center) is bound between the two subunits at the base of the
catcher's mitts in the unswapped conformation (B). Two molecules
of H[4]B (yellow, center, on edge) are also bound at the
interface and line the active-center channels leading to the
hemes.
|
|
Figure 5.
Figure 5 Potential interaction surfaces of iNOS[ox]. (A and B)
Electrostatic potential mapped onto the solvent-accessible
molecular surface of the unswapped zinc-bound iNOS[ox] dimer. In
the left orientation (A) (matching Figure 1), surface
surrounding the exposed heme edge (Region 1) is surrounded by
significant positive (blue) electrostatic potential (contoured
at 3 kT/q; k = Boltzmann constant, T = temperature, q = 1 point
charge), whereas the region surrounding the zinc site [(B),
Region 2] (right view, rotated 90° about a horizontal axis) is
neutral or negative (red). A pocket adjoining Region 1 and near
the heme-ligating thiolate also has significant positive
potential and residue conservation (Region 3). (C and D)
Solvent-accessible surface of the iNOS[ox] dimer (one subunit
red, the other subunit blue) color coded by residue conservation
(paler to more saturated represents less conserved to more
conserved), based on a group of NOS oxygenase domain sequences
representative of known species and isozymes. Conservation of
surface residues is most pronounced around the exposed heme edge
(Region 1) and in a region proximal to the heme thiolate (Region
3), and is low around the zinc site (Region 2).
|
|
|
|
|
|
The above figures are
reprinted
from an Open Access publication published by Macmillan Publishers Ltd:
EMBO J
(1999,
18,
6271-6281)
copyright 1999.
|
|
|
Secondary reference #1
|
|
|
Title
|
|
Structure of nitric oxide synthase oxygenase dimer with pterin and substrate.
|
|
|
Authors
|
|
B.R.Crane,
A.S.Arvai,
D.K.Ghosh,
C.Wu,
E.D.Getzoff,
D.J.Stuehr,
J.A.Tainer.
|
|
|
Ref.
|
|
Science, 1998,
279,
2121-2126.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Figure 1.
Fig. 1. NOS[ox]  -  fold, dimer
assembly, and likely interaction surface for NOS[red] and
caveolin. (A) The symmetric iNOS[ox] dimer viewed along the
crystallographic twofold axis, showing left (and^ right)
subunits with orange (yellow) winged sheets and
flanking blue (cyan) helices. Ball-and-stick models (white bonds
with red^ oxygen, blue nitrogen, yellow sulfur, and purple iron
atoms) highlight active-center hemes (left-most and right-most),
interchain disulfide^ bonds (center, foreground), pterin
cofactors (white, left-center and right-center), and substrate
L-Arg (green left and magenta^ right). The NH[2]-terminal ends
contribute hairpins
(center top and bottom) to the dimer interface, and the
COOH-termini (lower left and upper right) lie 85 Å apart.
Gray loops (residues 101^ to 107) are disordered. (B) iNOS[ox]
dimer shown rotated^ 90° about a horizontal axis from (A).
Each heme is cupped between the inward-facing palm (webbed sheet) and
thumb (magenta loop in front of left heme and green loop behind
right heme) of the^ "catcher's mitt" subunit fold. (C)
Solvent-accessible surface^ (29) of the iNOS[ox] dimer (one
subunit red, one subunit blue) oriented as in (B) and
color-coded by residue conservation (paler to more saturated
represents less conserved to more conserved) in NOS[ox]
sequences of known species and isozymes. The heme (white^ tubes)
is also solvent-exposed on the side (left subunit) opposite^ the
active-center channel (right subunit) and surrounded by a^
highly conserved hydrophobic surface for NOS[red] and caveolin
binding. (Stereo variations of Figs.
|
|
Figure 5.
Fig. 5. Proposed L-Arg-assisted NOS oxygen activation. First,
substrate L-Arg (only guanidinium shown) donates a proton to
peroxo-iron, facilitating O-O bond cleavage and conversion to a
proposed oxo-iron(IV)  -cation
radical species, which then rapidly hydroxylates the^ neutral
guanidinium to NOH-L-Arg, possibly through a radical-based^
mechanism (3).
|
|
|
|
|
|
The above figures are
reproduced from the cited reference
with permission from the AAAs
|
|
|
|
|
|
|
