Chemistry Reference
In-Depth Information
membrane), cytochrome P450 macromolecules are associated with another protein,
NADPH/cytochrome P450 reductase. The latter enzyme is converted to its reduced
form by the action of NADPH (reduced form of nicotine adenine dinucleotide phos-
phate). Electrons are passed from the reduced reductase to cytochrome P450, con-
verting it to the Fe
2+
state.
Xenobiotic substrates attach themselves to the hydrophobic binding site of P450,
when the iron of the hemeprotein is in the Fe
3+
state. After a single electron has been
passed from the reductase to P450, the hemeprotein moves into the Fe
2+
state, and
molecular oxygen can now bind to the enzyme:substrate complex. It binds to the free
sixth ligand position of the iron, where it is now in close proximity to the bound lipo-
philic substrate (Figure 2.3). A further electron is then passed to P450, and this leads
to the activation of the bound oxygen. This second electron may come from the same
source as the first, or it may originate from another microsomal hemeprotein, cyto-
chrome b5, which is reduced by NADH rather than NADPH. After this, molecular
oxygen is split—one atom being incorporated into the xenobiotic metabolite, and the
other into water. The exact mechanism involved in these changes is still controver-
sial. However, a widely accepted version of the main events is shown in Figure 2.4.
The uptake of the second electron leads to the formation of a highly reactive super-
oxide anion, O
2
−
, after which the splitting of molecular oxygen and “mixed function
oxidation” immediately follow. The P450 returns to the Fe
3+
state, and the whole
cycle can begin again.
“Active” oxygen generated at the catalytic center of cytochrome P450 can attack
the great majority of organic molecules that become attached to the neighboring
substrate-binding site (Figure 2.3). When substrates are bound, the position of the
molecule that is attacked (“regioselectivity”) will depend on the spatial relationship
between the bound molecule and the activated oxygen. Active oxygen forms are most
likely to attack the accessible positions on the xenobiotic which are nearest to them.
Differences in substrate specificity between the many different P450 forms are due,
O
2
e
-
(
from NADPH
)
XH - Fe
2+
(XH - Fe(II) O
2
)
2+
XH - Fe
3+
e
-
(
from NADPH
or NADH
)
XH
(XH - Fe(II)O
2
-
)
+
Fe
3+
XOH
2H
+
(XH - Fe(III)O)
3+
H
2
O
fIgure 2.4
Proposed mechanism for monooxygenation by cytochrome P450.
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