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Evolution of acyl-ACP thioesterases and β-ketoacyl-ACP synthases revealed by protein–protein interactions

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Abstract

The fatty acid synthase (FAS) is a conserved primary metabolic enzyme complex capable of tolerating cross-species engineering of domains for the development of modified and overproduced fatty acids. In eukaryotes, acyl-acyl carrier protein thioesterases (TEs) off-load mature cargo from the acyl carrier protein (ACP), and plants have developed TEs for short/medium-chain fatty acids. We showed that engineering plant TEs into the green microalga Chlamydomonas reinhardtii does not result in the predicted shift in fatty acid profile. Since fatty acid biosynthesis relies on substrate recognition and protein–protein interactions between the ACP and its partner enzymes, we hypothesized that plant TEs and algal ACP do not functionally interact. Phylogenetic analysis revealed major evolutionary differences between FAS enzymes, including TEs and ketoacyl synthases (KSs), in which the former is present only in some species, whereas the latter is present in all, and has a common ancestor. In line with these results, TEs appeared to be selective towards their ACP partners, whereas KSs showed promiscuous behavior across bacterial, plant, and algal species. Based on phylogenetic analyses, in silico docking, in vitro mechanistic cross-linking, and in vivo algal engineering, we propose that phylogeny can predict effective interactions between ACPs and partner enzymes.

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Acknowledgments

Strain CY1877 was a generous gift of John E. Cronan (University of Illinois). The plasmid harboring VhACP was a generous gift of Peter Murphy and David Byers (Dalhousie University). This work was supported by California Energy Commission CILMSF 500-10-039; Department of Energy DE-EE0003373; National Institute of Health R01GM094924 and R01GM095970.

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Authors and Affiliations

  1. Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA, 92093-0358, USA

    Joris Beld, Jillian L. Blatti & Michael D. Burkart

  2. Sapphire Energy, Inc., San Diego, CA, USA

    Craig Behnke & Michael Mendez

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  1. Joris Beld
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  2. Jillian L. Blatti
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  3. Craig Behnke
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  4. Michael Mendez
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  5. Michael D. Burkart
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Correspondence to Michael D. Burkart.

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Joris Beld and Jillian L. Blatti contributed equally to this work.

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Beld, J., Blatti, J.L., Behnke, C. et al. Evolution of acyl-ACP thioesterases and β-ketoacyl-ACP synthases revealed by protein–protein interactions. J Appl Phycol 26, 1619–1629 (2014). https://doi.org/10.1007/s10811-013-0203-4

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