Biology Reference
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due to rapid autoxidation and/or low affinity for O
2
(
Tanaka, Takahashi, &
Shimizu, 2007
). The rotation of the propionate groups of the haem also
occurs upon O
2
binding, which may trigger the formation of the hydrogen
bond between Asn84 and Tyr126 through the haem propionate 6
hydrogen-bond network (
El-Mashtoly, Takahashi, Shimizu, & Kitagawa,
2007; Sato et al., 2002
). These hydrogen-bond networks are proposed to
be responsible for the intramolecular signal transduction from the sensor
to enzymatic domains (
El-Mashtoly et al., 2007
).
5.2.4 Regulation of PDEs activity of EcDos
The EAL domain, which contains a conserved sequence motif of Glu-Ala-
Leu as the enzymatic active site, shows PDE activity with a substrate of cyclic
di-GMP (
Delgado-Nixon et al., 2000; Tuckerman et al., 2009
). The phys-
iological function of
Ec
Dos is regulated by exogenous ligand binding to the
haem. Thus, the PDE activity is enhanced upon O
2
binding to the Fe
2
รพ
haem (
Tanaka et al., 2007; Tuckerman et al., 2009
). CO and NO binding
to the haem causes similar enhancement of the PDE activity (
Tanaka et al.,
2007; Tuckerman et al., 2009
), indicating that
Ec
Dos cannot discriminate
these gas molecules. Positive cooperativity in O
2
binding is observed for
Ec
Dos (
Lechauve et al., 2009; Tuckerman et al., 2009
).
Lechauve et al.
(2009)
have reported that the second O
2
molecule binds to
Ec
Dos with a
sixfold higher intrinsic affinity than the first one. They propose that the
replacement of Met95 by O
2
and its swapping out of the haem pocket in
one subunit leads to a modification at the dimer interface, presumably lead-
ing to decrease Met95 affinity for haem, which will increase O
2
-binding
affinity, in the other subunit. The PDE activity also shows a non-linear
dependence on the fraction of the O
2
-saturated haem, which enables to acti-
vate
Ec
Dos in narrow range of O
2
concentrations (
Kobayashi et al., 2010;
Tuckerman et al., 2009
).
The basal activity of the ferrous
Ec
Dos increases to a similar value for O
2
-,
CO-, or NO-bound form by mutation of Met95 to Ala or Leu (
Tanaka et al.,
2007
). These results suggest that the dissociation of Fe-Met bond upon ligand
binding results in a conformational change of the FG loop to activate the PDE
activity of
Ec
Dos. In addition to Met95, several amino acid residues are iden-
tified by mutagenesis and resonance Raman studies as those responsible for
controlling ligand-binding properties and/or enzymatic activity (
El-
Mashtoly, Nakashima, Tanaka, Shimizu, & Kitagawa, 2008; Ishitsuka et al.,
2008; Ito, Araki, et al., 2009; Ito, Igarashi, & Shimizu, 2009; Tanaka &
Shimizu, 2008
).
