Chemistry Reference
In-Depth Information
O
O
O
HMGS
O
O
OH
+
CoA
CoA
CoA
S
S
O
S
acetyl-CoA
acetoacetyl-CoA
3-hydroxyl-3-methylglutaryl-CoA
(a)
KS
C?S
O
O
O
AT
HS
ACP
O
S
S
ACP
ACP
O
O
O
O
O
R
OH
HMGS
+
O
S
S
ACP
R
S
ACP
ACP
H
2
R
O
S
ACP
O
R
O
ECH
ECH
1
2
R
O
O
S
ACP
S
ACP
H
2
CO
2
(b)
Figure 7.5
HMGS cassette reaction scheme. (a) HMG-CoA synthase (HMGS) reaction
from primary metabolism. (b) An HMGS cassette can convert the
β
-ketone to an alkene
(
β
,
γ
or
γ
,
δ
double bond) with a pendant methyl (or ethyl) group.
the formation of the HMG-intermediate is the generation of of acetyl-ACP. This
step is accomplished through the loading via an AT of malonyl-CoA [or perhaps
methylmalonyl-CoA in at the case of TaE (58)]. Then, the decarboxylative KS
converts the malonyl-ACP into acetyl-ACP, after which the tethered acetyl group
is condensed onto the
β
-ketone of the polyketide intermediate. Finally, formation
of the HMG-analog is completed on addition of water.
Processing of the HMG-intermediate can vary considerably, but typically
proceeds via dehydration and decarboxylation catalyzed by two enoyl-CoA
hydratase-like domains (Fig. 7.5b). Based on sequence similarity, the members
of the crotonase fold family observed in these HMGS cassettes can be subdivided
into two groups, termed ECH
1
and ECH
2
(54). The successive dehydration and
decarboxylation steps are catalyzed by the ECH
1
and ECH
2
enzymes/domains,
respectively. Evidence for the specific function of the curacin ECH
1
and ECH
2
enzyme pair from the curacin pathway has been demonstrated using a coupled
enzyme assay and ESI-FT-ICR MS (54). Using purified ECH
1
(CurE) and ECH
2
(the N-terminal domain of CurF) overexpressed in
E. coli
,(
S
)-HMG-ACP was
