Chemistry Reference
In-Depth Information
studies suggested that the formation of the terminal double bond develops from
successive decarboxylation and dehydration events (35). The biosynthetic gene
cluster responsible for curacin A biosynthesis has been identified and initially
characterized, which enables the putative assignment of domains within the pre-
dicted elongation modules (35). Like most other known polyketide biosynthetic
pathways, the final elongation module of the curacin pathway, CurM, contains
a terminal thioesterase domain that presumably plays a role in formation of the
terminal olefin; however, biochemical evidence for such a role is lacking. Inter-
estingly, domain analysis of CurM also predicts the presence of a sulfotransferase
(ST) domain immediately preceding the TE domain. Typically, ST domains are
responsible for transferring a sulfuryl group from a donor molecule (such as
3
-phosphoadenosine-5
-phosphosulfate, PAPS) to a variety of acceptor carbo-
hydrates, proteins and other low-molecular weight metabolites (38). Although
STs have been characterized from both eubacterial and eukaryotic organisms, the
presence of an ST domain within a PKS system is unprecedented.
Current efforts in our laboratory include elucidating the roles of the ST and
TE domains in curacin A biosynthesis, and in particular, their potential functions
in terminal olefin formation. One possible mechanism that is currently under
consideration is shown in Fig. 7.3b. On reduction of the
β
-keto functionality of
the ACP-bound thioester intermediate, the ST domain transfers a sulfuryl group
from the donor molecule PAPS to the 3-hydroxyl group of the thioester chain.
Consistent with this hypothesis, bioinformatic analysis suggests that the putative
curacin ST domain contains the signature PAPS binding pocket (unpublished
data); however, no experimental evidence suggests that this ST domain can cat-
alyze the sulfuryl transfer or that the ST domain can bind the requisite PAPS
donor molecule. Assuming our hypothesis is correct and that the ST domain
functions as proposed, transfer of the sulfurylated intermediate to the TE domain
would initiate hydrolytic termination of curacin A biosynthesis to produce the
linear free acid. At this point, one of several chemical steps can be envisioned.
Following hydrolysis, the TE may catalyze decarboxylation, after which the for-
mation of the double bond would occur in a concerted process by displacement
of the sulfate leaving group. Alternatively, a separately encoded enzyme might
be responsible for decarboxylating the free acid generated by the TE domain. It
is also conceivable that on TE catalyzed hydrolysis, the decarboxylation reaction
occurs spontaneously due to the presence of the sulfate leaving group at carbon 3.
Similar to curacin, the polyketide metabolite tautomycetin
14
also possesses a
terminal olefin. This polyketide metabolite has potential medicinal value because
of its novel immunosuppressive activities (39). The tautomycetin biosynthetic
gene cluster has been sequenced recently, which enables domain composition
analysis of the terminal elongation module (40). Unlike the terminal module
involved in curacin biosynthesis, the final tautomycetin elongation module does
not contain the unusual ST domain. This finding may suggest that formation of
the terminal olefin of tautomycetin
14
occurs by a different chemical mechanism.
The final elongation module does contain a TE domain, which presumably termi-
nates tautomycetin biosynthesis through generation of the free acid (Fig. 7.3c).
