Pleuromutilin belongs to the class of secondary metabolites known as
terpenes, which are produced in fungi through the
mevalonate pathway (MEP pathway). Its synthetic bottleneck lays on the production of the precursor
GGPP and the following formation of the tricyclic structure, which is catalyzed by
Pl-cyc, a bifunctional diterpene synthase (DTS). This Cyclase shows a new class II DTS activity, catalyzes a ring contraction and the formation of a 5-6-bicyclic ring structure. Specifically, DTS shows two catalytic distinguishable domains: On the one hand it has at the N-terminal region a class II DTS domain, which catalyzes a cascade cyclization, resulting in a
decalin core. Subsequently, variable 1,2-proton and methyl shifts occur to translocate the
carbocation towards one of the two interconnecting C-atoms and this intermediate induces a base-catalyzed ring contraction. Therefore, class II DTS promotes in general a ring contraction during the cyclisation of
GGPP. On the other hand, at the C-terminal end it has a class I DTS domain, which catalyzes a conjugated dephosphorylation, generating the 8-membered cyclic core, followed by a 1,5-proton shift and a stereospecific hydroxylation to obtain premutilin. (GGPP). The following cyclisation steps to the pleuromutilin tricyclic core will be provided by class II and class I terpene synthase domain of
Pl-cyc. Final catalytic, stereospecific
hydroxylation at C-11 and C-3 (
Pl-p450-1, Pl-p450-2), regiospecific oxidation of the 3-hydroxy group to a
ketone (
Pl-sdr), acetyl-group-transfer at OH-14 (
Pl-atf) and final hydroxylation at the α-acetyl position (
Pl-p450-3) will lead to pleuromutilin. The P450-1 and P450-2 are essential for hydroxylation of two ring structures regarding the premutilin skeleton, oxidating specifically at position C-11 and C-3, respectively. The
short-chain dehydrogenase/reductase enzyme (
Pl-sdr) has a regiospecific activity and converts the 3-hydroxy group to a ketone, forming the intermediate
mutilin. Acetyltransferase (
Pl-atf) catalyzes the transfer of acetyl group to 14-OH of mutilin. Finally,
Pl-p450-3 hydroxylates the α-methyl group of the acetyl side chain generating pleuromutilin. == References ==