The
neuro
nal isoform of
nitric oxide sy
nthase (
nNOS), the e
nzyme respo
nsible for the productio
n of
nitricoxide i
n the ce
ntral
nervous system, represe
nts a
n attractive target for the treatme
nt of various
neurodege
nerative disorders. X-ray crystal structures of complexes of
nNOS with two
nNOS-selectivei
nhibitors, (4
S)-
N-{4-ami
no-5-[(2-ami
noethylami
no]pe
ntyl}-
N'-
nitrogua
nidi
ne (
1) a
nd 4-
N-(
N![](/images/gifchars/omega.gif)
-
nitro-
L-argi
ni
nyl)-
trans-4-ami
no-
L-proli
ne amide (
2), led to the discovery of a co
nserved structural water moleculethat was hydroge
n bo
nded betwee
n the two heme propio
nates a
nd the i
nhibitors (Figure 2). O
n the basis ofthis observatio
n, we hypothesized that by attachi
ng a hydroge
n bo
nd do
nor group to the amide
nitroge
n of
2 or to the seco
ndary ami
ne
nitroge
n of
1, the i
nhibitor molecules could displace the structural water moleculea
nd obtai
n a direct i
nteractio
n with the heme cofactor. To test this hypothesis, peptidomimetic a
nalogues
3-5, which have either a
n N-hydroxyl (
3 a
nd
5) or
N-ami
no (
4) do
nor group, were desig
ned a
nd sy
nthesized.X-ray crystal structures of
nNOS with i
nhibitors
3 a
nd
5 bou
nd verified that the
N-hydroxyl group had,i
ndeed, displaced the structural water molecule a
nd provided a direct i
nteractio
n with the heme propio
natemoiety (Figures 5 a
nd 6). Surprisi
ngly, i
n vitro activity assay results i
ndicated that the additio
n of a hydroxylgroup (
3) o
nly i
ncreased the pote
ncy slightly agai
nst the
neuro
nal isoform over the pare
nt compou
nd (
1).Ratio
nalizatio
ns for the small i
ncrease i
n pote
ncy are co
nsiste
nt with other cha
nges i
n the crystal structures.