Engineered Chimeras Unveil Swappable Modular Features of Fatty Acid and Polyketide Synthase Acyl Carrier Proteins

Biochemistry. 2022 Feb 15;61(4):217-227. doi: 10.1021/acs.biochem.1c00798. Epub 2022 Jan 24.


The strategic redesign of microbial biosynthetic pathways is a compelling route to access molecules of diverse structure and function in a potentially environmentally sustainable fashion. The promise of this approach hinges on an improved understanding of acyl carrier proteins (ACPs), which serve as central hubs in biosynthetic pathways. These small, flexible proteins mediate the transport of molecular building blocks and intermediates to enzymatic partners that extend and tailor the growing natural products. Past combinatorial biosynthesis efforts have failed due to incompatible ACP-enzyme pairings. Herein, we report the design of chimeric ACPs with features of the actinorhodin polyketide synthase ACP (ACT) and of the Escherichia coli fatty acid synthase (FAS) ACP (AcpP). We evaluate the ability of the chimeric ACPs to interact with the E. coli FAS ketosynthase FabF, which represents an interaction essential to building the carbon backbone of the synthase molecular output. Given that AcpP interacts with FabF but ACT does not, we sought to exchange modular features of ACT with AcpP to confer functionality with FabF. The interactions of chimeric ACPs with FabF were interrogated using sedimentation velocity experiments, surface plasmon resonance analyses, mechanism-based cross-linking assays, and molecular dynamics simulations. Results suggest that the residues guiding AcpP-FabF compatibility and ACT-FabF incompatibility may reside in the loop I, α-helix II region. These findings can inform the development of strategic secondary element swaps that expand the enzyme compatibility of ACPs across systems and therefore represent a critical step toward the strategic engineering of "un-natural" natural products.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acyl Carrier Protein / chemistry
  • Acyl Carrier Protein / metabolism*
  • Amino Acid Sequence
  • Chimera / metabolism
  • Escherichia coli / enzymology
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism*
  • Fatty Acid Synthase, Type II / metabolism
  • Fatty Acid Synthases / chemistry
  • Fatty Acid Synthases / metabolism*
  • Fatty Acids / metabolism
  • Molecular Dynamics Simulation
  • Polyketide Synthases / chemistry
  • Polyketide Synthases / metabolism*
  • Polyketides / metabolism
  • Surface Plasmon Resonance / methods
  • Transferases (Other Substituted Phosphate Groups) / metabolism


  • Acyl Carrier Protein
  • Escherichia coli Proteins
  • Fatty Acids
  • Polyketides
  • Polyketide Synthases
  • polyketide beta-ketoacylsynthase
  • Fatty Acid Synthases
  • Transferases (Other Substituted Phosphate Groups)
  • holo-(acyl-carrier-protein) synthase
  • Fatty Acid Synthase, Type II