Unusual Type II Polyketide Synthase System Involved in Cinnamoyl Lipid Biosynthesis.

As a unique structural moiety in natural products, cinnamoyl lipid (CL), was proposed to be assembled via unusual type II polyketide synthases (PKSs). However, their biosynthetic machinery remains to be disclosed. Herein, we demonstrated the assembly line of CL compounds, youssoufenes, which is accomplished by a PKS system uniquely harbouring three phylogenetically different ketosynthase/chain length factor  (KS/CLF) complexes (YsfB/C, YsfD/E, and YsfJ/K). Through in vivo gene inactivation, in vitro reconstitution, as well as intracellular tagged carrier-protein tracking (ITCT) strategy developed in this study, we successfully captured the isomerase-dependent ACP-tethered polyunsaturated chain elongation process. The three KS/CLFs were unravelled to modularly assemble different parts of the youssoufene skeleton, during which benzene ring closure happens right after ACP-tethered C18 polyene formation. Of note, the ITCT strategy would significantly contribute to disclosing the mysteries of carrier proteins-dependent biosynthetic machineries.