Introduction to Enzyme and Coenzyme Chemistry - E-Library Home

Enzymatic Carbon–Carbon Bond Formation 175
found to act as a catalyst for the Dieckmann condensation of diethyl adipate,
generating an epoxide byproduct. A mechanism was proposed for this reaction
in which the naphthoxide salt and dioxygen react via a dioxetane intermediate
to form a tertiary alkoxide species which acts as a base for the ester condensation
reaction. Note that the naphthoxide salt itself is not a strong enough base
to catalyse the reaction, but is converted into a much stronger base via reaction
with oxygen.
In order to investigate the mechanism of the enzyme-catalysed carboxylation
reaction, reduced vitamin K was incubated with carbon dioxide and
enzyme in the presence of 18 O 2 . Complete incorporation of 18 O into the epoxide
oxygen of vitamin K epoxide was observed, together with partial incorporation
of 18 O (approx. 20%) into the ketone carbonyl, consistent with the dioxetane
mechanism. It has been proposed that the hydrate alkoxide ion released from
fragmentation of the dioxetane intermediate acts as a base to deprotonate the
g-H of a glutamyl substrate, followed by reaction with carbon dioxide, as
shown in Figure 7.24. It has been shown that the C-4 proS hydrogen is
abstracted, and subsequent reaction with carbon dioxide occurs with inversion
of conWguration. Note that it is remarkable that the hydrate alkoxide ion does
not simply collapse to form the corresponding ketone: presumably the microenvironment
of the enzyme active site disfavours the loss of hydroxide ion from
the lower face of vitamin K.
O −
R
O 2
O
R
O −
R
OH
HO O O −
HO
O O
O
R
O
O
R
O
O
R
O
O
CO 2
−
CO 2
−
HO OH
− O 2 C
H
−
O
C
O
HO O −
H S CO −
2
H R
N
H
O
N
H
O
N
H
O
Figure 7.24 Mechanism for the vitamin K-dependent carboxylase.