Substrate Controlled Divergence in Polyketide Synthase Catalysis

Biochemical
characterization of polyketide synthases (PKSs) has
relied on synthetic substrates functionalized as electrophilic esters
to acylate the enzyme and initiate the catalytic cycle. In these efforts, N-acetylcysteamine thioesters have typically been employed
for in vitro studies of full PKS modules as well as excised domains.
However, substrate engineering approaches to control the catalytic
cycle of a full PKS module harboring multiple domains remain underexplored.
This study examines a series of alternatively activated native hexaketide
substrates on the catalytic outcome of PikAIV, the sixth and final
module of the pikromycin (Pik) pathway. We demonstrate the ability
to control product formation with greater than 10:1 selectivity for
either full module catalysis, leading to a 14-membered macrolactone,
or direct cyclization to a 12-membered ring. This outcome was achieved
through modifying the type of hexaketide ester employed, demonstrating
the utility of substrate engineering in PKS functional studies and
biocatalysis.