Discovery and Engineering of Pathways for Production of α-Branched Organic Acids

Michael R. Blaisse, Hongjun Dong, Beverly Fu, Michelle C.Y. Chang

Research output: Contribution to journalArticlepeer-review

14 Scopus citations


Cell-based synthesis offers many opportunities for preparing small molecules from simple renewable carbon sources by telescoping multiple reactions into a single fermentation step. One challenge in this area is the development of enzymatic carbon-carbon bond forming cycles that enable a modular disconnection of a target structure into cellular building blocks. In this regard, synthetic pathways based on thiolase enzymes to catalyze the initial carbon-carbon bond forming step between acyl coenzyme A (CoA) substrates offer a versatile route for biological synthesis, but the substrate diversity of such pathways is currently limited. In this report, we describe the identification and biochemical characterization of a thiolase-ketoreductase pair involved in production of branched acids in the roundworm, Ascaris suum, that demonstrates selectivity for forming products with an α-methyl branch using a propionyl-CoA extender unit. Engineering synthetic pathways for production of α-methyl acids in Escherichia coli using these enzymes allows the construction of microbial strains that produce either chiral 2-methyl-3-hydroxy acids (1.1 ± 0.2 g L-1) or branched enoic acids (1.12 ± 0.06 g L-1) in the presence of a dehydratase at 44% and 87% yield of fed propionate, respectively. In vitro characterization along with in vivo analysis indicates that the ketoreductase is the key driver for selectivity, forming predominantly α-branched products even when paired with a thiolase that highly prefers unbranched linear products. Our results expand the utility of thiolase-based pathways and provide biosynthetic access to α-branched compounds as precursors for polymers and other chemicals.

Original languageEnglish (US)
Pages (from-to)14526-14532
Number of pages7
JournalJournal of the American Chemical Society
Issue number41
StatePublished - Oct 18 2017
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Catalysis
  • General Chemistry
  • Biochemistry
  • Colloid and Surface Chemistry


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