Repurposing a Fully\nReducing Polyketide Synthase toward\n2‑Methyl Guerbet-like Lipids

In nature, thousands of diverse and bioactive polyketides\nare assembled\nby a family of multifunctional, “assembly line” enzyme\ncomplexes called polyketide synthases (PKS). Since the late 20th century,\nthere have been several attempts to decode, rearrange, and “reprogram”\nthe PKS assembly line to generate valuable materials such as biofuels\nand platform chemicals. Here, the first module from Mycobacterium tuberculosis (<i>Mt</i>)\nPKS12, an unorthodox, “modularly iterative” PKS, was\nmodified and repurposed toward the formation of 2-methyl Guerbet lipids,\nwhich have wide applications in industry. We established a robust\nmethod for the recombinant expression and purification of this modified\nmodule (named [M<sub>1</sub>*]), and we demonstrated its ability to\ncatalyze the formation of several 2-methyl Guerbet-like lipids (C<sub>13</sub>–C<sub>21</sub>). Furthermore, we studied and applied\nthe promiscuous thioesterase activity of a neighboring β-ketoacyl\nsynthase (KS) to release [M<sub>1</sub>*]-bound condensation products\nin a one-pot biosynthetic cascade. Finally, starting from lauric acid,\nwe could generate our primary target compound (2-methyltetradecanoic\nacid) by coupling the Escherichia coli fatty acyl-CoA synthetase FadD to [M<sub>1</sub>*]. This work supports\nthe biosynthetic utility of engineered PKS modules such as [M<sub>1</sub>*] and their ability to derive valuable Guerbet-like lipids\nfrom inexpensive fatty acids.