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Enzymatic Redox Chemistry 131
R
N
H
N
O
R H +
N N
O
R
N
N
O
N
H
O
NH
N
H
O
NH
N
H
O
NH
1-electron transfer to O 2 O 2
O 2
recombination
R
N
N
O
R
N
N
O
NH
N
O
HO −
2
Figure 6.14 Regeneration of oxidised Xavin by molecular oxygen.
N
H
O
O
HO
NH
substrate to oxidised Xavin (a); nucleophilic attack by the substrate on the Xavin
nucleus, either by a heteroatom at C-4a (b), or by a substrate carbanion at N-5
(c); radical mechanisms involving single electron transfer (d).
Nucleophilic attack on the Xavin nucleus (Figure 6.15b) is well precedented
in Xavin model systems, but there is no Wrm evidence for the existence of
covalent substrate–Xavin adducts in Xavin-dependent dehydrogenases. We
shall examine the available evidence for the enzymes acyl coenzyme A (CoA)
dehydrogenase and monoamine oxidase.
There is good evidence in the reaction of acyl CoA dehydrogenase that the
Wrst step of the mechanism involves deprotonation of the substrate adjacent to
the thioester carbonyl, since the enzyme catalyses the exchange of a C a proton
with 2 H 2 O. However, there is also evidence for an a-radical intermediate, since
the enzyme is inactivated by a substrate analogue containing a b-cyclopropyl
group. This inhibitor is a metabolite of hypoglycin A, which is the causative
agent of Jamaican vomiting sickness. The proposed mechanism of inactivation
is shown in Figure 6.16. Upon formation of the a-radical a rapid opening of the
strained cyclopropyl ring takes place, giving a rearranged radical intermediate
which recombines with the Xavin semiquinone irreversibly.
A possible mechanism for acyl CoA dehydrogenase consistent with both
observations is shown in Figure 6.17. Deprotonation to form an a-carbanion is
followed by a one-electron transfer from the substrate to FAD, generating a
substrate radical and the Xavin semiquinone. The reaction can be completed by
abstraction of H .
at the b-position by the Xavin radical, generating reduced
FADH 2 . In support of the Wnal proposed step in the mechanism there are
isolated examples of Xavo-enzymes which catalyse hydrogen atom transfer
from FADH 2 to the b-position of a,b-unsaturated carbonyl substrates (see
Problem 4).