Chemistry Reference
In-Depth Information
FIGURE 13.10
The catalytic cycle of CytcO. The superscript indicates the number of electrons transferred to the catalytic site. The electrons
(e
) and protons at the arrows (in green) are those transferred to the catalytic site, while the protons indicated by arrows (in red) perpendicular
to the reaction arrows indicate pumped protons. Y is Tyr288 (see text) where Y*O indicates a tyrosyl radical. The oxidized state is O
4(0)
(see
text). When both haem a
3
and Cu
B
are reduced (R
2
), O
2
binds to form state A
2
after which the reaction proceeds as described in the text. The
pathway along the black arrows indicates the reaction during turnover of the enzyme when electrons are added one by one to the catalytic site.
The reaction pathway along the blue arrows is that observed during reaction of the fully reduced CytcO (with four electrons) with O
2
. Here,
after binding of O
2
to haem a
3
(to form state A
2
), an electron is transferred to the catalytic site (to form P
3
) before the proton is transferred to the
catalytic site (to form F
3
). Note that in state P
3
there is an excess negative charge as compared to the other states.
(From
Brzezinski &
Johansson, 2010
.
Copyright 2011, with permission from Elsevier.)
is not linked to any proton uptake from solution and, protons and electrons are only relocated locally within the
catalytic site resulting in oxidation of haem a
3
and Cu
B
. In addition, one electron (and a proton) is transferred from
residue Tyr288, located within the catalytic site, which forms a tyrosyl radical, Tyr288$, and structural changes
occur in the vicinity of the catalytic site. In the next step an electron is transferred to the Tyr radical, accompanied
by proton uptake from the n-side to form state F
3
and proton pumping with a time constant of 100
m
s. In the final
step, with a time constant of
1.2 ms, the last electron is transferred to the catalytic site, also accompanied by
proton uptake from the n-side forming state O
4
and proton pumping. When considering the oxidation state of the
catalytic site, the O
4
state is equivalent to O
0
as the enzyme becomes fully oxidized when four electrons have been
transferred to O
2
.
Part of the energy released in the redox reaction is conserved by vectorial transfer of protons across the
membrane from the N-side to the P side, thereby maintaining an electrochemical proton gradient that is used for
the synthesis of ATP. Two proton transfer pathways leading from the N-side surface toward the binuclear centre
have been identified (
Figure 13.9
(
b)). In cytochrome c oxidase from Rhodobacter sphaeroides, one of the
pathways (D-pathway) starts with Asp132 and leads to Glu286. Since the D-pathway is used both for the substrate
protons, which are transferred to the catalytic site, and pumped protons, which are transferred to a proton-
accepting group in the exit pathway, there must be a branching point within the pathway from where protons can
be transferred either toward the dinuclear centre or toward the output side of the enzyme. This is thought to be at
Glu288. The other pathway (K-pathway) starts at the N-side surface at Glu101 and leads via a highly conserved
Lys362 and Tyr288 to the dinuclear centre.
Figure 13.9
(
b) also illustrates analogies between CcO and peroxidases and catalases, which we discuss next, in
terms of both oxygen
w
oxygen bond cleavage chemistry and the nature of the products of the reactions. In CcO, the
enzyme extracts three electrons from metals in the active site
e
e
two from haem a
3
as it goes from the
þ
2 to the
þ
4
state and one from Cu
B
as it is oxidized from cuprous to cupric
e
and one electron from a redox-active protein side
