Synthetic Polyketide Enzymology: Platform for Biosynthesis of Antimicrobial Polyketides

Abstract

Synthetic biology often employs enzymes in the biosynthesis of compounds for purposeful function. Here, we define synthetic enzymology as the application of enzymological principles in synthetic biology and describe its use as an enabling platform in synthetic biology for the purposeful production of compounds of biomedical and commercial importance. In particular, we demonstrated the use of synthetic polyketide enzymology as a means to develop lead polyketide based compounds for antimicrobial therapeutics, as exemplified by the modular coupling of acid:CoA ligases to type III polyketide synthases in the biosynthesis and development of polyketide-based biochemicals. Using wild-type and rationally designed mutants of a type III polyketide synthase isolated from Oryza sativa (OsPKS), we produced a chemically diverse library of novel polyketides and identified two bioactive antimicrobials, 4-hydroxy-6-[(1<i>E</i>)-2-(4-hydroxyphenyl)­ethenyl]-2<i>H</i>-pyran-2-one (bisnoryangonin) and 3,6,7-trihydroxy-2-(4-methoxybenzyl)-4<i>H</i>-1-benzopyran-4,5,8-trione (26OH), respectively, from a screen against a collection of Gram-positive and Gram-negative bacteria. The purification, crystallization, and structural resolution of recombinant OsPKS at 1.93 Å resolution are also reported. Using the described route of synthetic polyketide enzymology, a library of OsPKS mutants was generated as an additional means to increase the diversity of the polyketide product library. We expect the utility of synthetic enzymology to be extended to other classes of biomolecules and translated to various purposeful functions as the field of synthetic biology progresses