(729b) A Polyketide Macrolactone Synthase from the Filamentous Fungus Gibberella Zeae

Abstract

Resorcylic
acid lactones represent a unique class of fungal polyketides and display a wide
range of biological activities, such as nanomolar inhibitors of Hsp90 and MAP
kinase.  The biosynthesis of these compounds is proposed to involve two fungal
polyketide synthases (PKS) that function collaboratively to yield a 14-membered
macrolactone with a resorcylate core.  We report here the reconstitution of Gibberella
zeae PKS13, which is the nonreducing PKS associated with zearalenone
biosynthesis. Using a small molecule mimic of the natural hexaketide starter
unit, we reconstituted the entire repertoire of PKS13 activities in vitro,
including starter unit selection, iterative condensation, regioselective C2-C7
cyclization and macrolactone formation.  PKS13 synthesized both natural
14-membered and novel 16-membered resorcylic acid lactones, indicating relaxed
control in both iterative elongation and macrocyclization.  Furthermore, PKS13
exhibited broad starter-unit specificities towards fatty acyl-CoAs ranging in
sizes between C6 and C16.  PKS13 was also proven to be able to interact with Escherichia
coli fatty acid biosynthetic machinery and can be primed with fatty-acyl
ACPp at low micromolar concentrations. A number of alkyl pyrones and alkyl
resorcylic esters were produced by E. coli BL21(DE3) harboring PKS13
with a highest yield up to 5 mg/L.  And we further demonstrated that BL21(DE3)
harboring PKS13 can synthesize new polyketides at a yield of 2~3 mg/L, when
supplemented with synthetic precursors, which showcases the utility of PKS13 in
precursor-directed biosynthesis. PKS13 is therefore a highly versatile
polyketide macrolactone synthase that is useful in the engineered biosynthesis
of novel polyketides, including resorcylic acid lactones that are not found in
nature.