Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Conformational flexibility of metazoan fatty acid synthase enables catalysis

Nature Structural & Molecular Biology volume 16, pages 190–197 (2009)Cite this article

Abstract

The metazoan cytosolic fatty acid synthase (FAS) contains all of the enzymes required for de novo fatty acid biosynthesis covalently linked around two reaction chambers. Although the three-dimensional architecture of FAS has been mostly defined, it is unclear how reaction intermediates can transfer between distant catalytic domains. Using single-particle EM, we have identified a near continuum of conformations consistent with a remarkable flexibility of FAS. The distribution of conformations was influenced by the presence of substrates and altered by different catalytic mutations, suggesting a direct correlation between conformation and specific enzymatic activities. We interpreted three-dimensional reconstructions by docking high-resolution structures of individual domains, and they show that the substrate-loading and condensation domains dramatically swing and swivel to access substrates within either reaction chamber. Concomitant rearrangement of the β-carbon–processing domains synchronizes acyl chain reduction in one chamber with acyl chain elongation in the other.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Reaction cycle in fatty acid biosynthesis.
Figure 2: Structural and functional organization of the metazoan FAS.
Figure 3: Conformational variability of Δ22-FAS in the absence of substrates.
Figure 4: Distribution of FAS conformations is altered in the presence of substrates.
Figure 5: Changes in domain position bring catalytic domains into proximity of the ACP to facilitate catalytic interactions.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Sul, H.S. & Smith, S. Fatty acid synthesis in eukaryotes. in Biochemistry of Lipids, Lipoproteins and Membranes (ed. Vance, D.E.a.V. J.E.) 155–190 (Elsevier, Amsterdam; Oxford, 2008).

    Chapter  Google Scholar 

  2. Kuhajda, F.P. et al. Fatty acid synthesis: a potential selective target for antineoplastic therapy. Proc. Natl. Acad. Sci. USA 91, 6379–6383 (1994).

    Article  CAS  Google Scholar 

  3. Loftus, T.M. et al. Reduced food intake and body weight in mice treated with fatty acid synthase inhibitors. Science 288, 2379–2381 (2000).

    Article  CAS  Google Scholar 

  4. Jenni, S. et al. Structure of fungal fatty acid synthase and implications for iterative substrate shuttling. Science 316, 254–261 (2007).

    Article  CAS  Google Scholar 

  5. Lomakin, I.B., Xiong, Y. & Steitz, T.A. The crystal structure of yeast fatty acid synthase, a cellular machine with eight active sites working together. Cell 129, 319–332 (2007).

    Article  Google Scholar 

  6. Asturias, F.J. et al. Structure and molecular organization of mammalian fatty acid synthase. Nat. Struct. Mol. Biol. 12, 225–232 (2005).

    Article  CAS  Google Scholar 

  7. Maier, T., Jenni, S. & Ban, N. Architecture of mammalian fatty acid synthase at 4.5 Å resolution. Science 311, 1258–1262 (2006).

    Article  CAS  Google Scholar 

  8. Rangan, V.S., Joshi, A.K. & Smith, S. Mapping the functional topology of the animal fatty acid synthase by mutant complementation in vitro. Biochemistry 40, 10792–10799 (2001).

    Article  CAS  Google Scholar 

  9. Witkowski, A. et al. Dibromopropanone cross-linking of the phosphopantetheine and active-site cysteine thiols of the animal fatty acid synthase can occur both inter- and intrasubunit. Reevaluation of the side-by-side, antiparallel subunit model. J. Biol. Chem. 274, 11557–11563 (1999).

    Article  CAS  Google Scholar 

  10. Burgess, S.A., Walker, M.L., Thirumurugan, K., Trinick, J. & Knight, P.J. Use of negative stain and single-particle image processing to explore dynamic properties of flexible macromolecules. J. Struct. Biol. 147, 247–258 (2004).

    Article  CAS  Google Scholar 

  11. Joshi, A.K., Witkowski, A., Berman, H.A., Zhang, L. & Smith, S. Effect of modification of the length and flexibility of the acyl carrier protein-thioesterase interdomain linker on functionality of the animal fatty acid synthase. Biochemistry 44, 4100–4107 (2005).

    Article  CAS  Google Scholar 

  12. Radermacher, M. The three-dimensional reconstruction of single particles from random and non-random tilt series. J. Electron Microsc. Tech. 9, 359–394 (1988).

    Article  CAS  Google Scholar 

  13. Keatinge-Clay, A.T. & Stroud, R.M. The structure of a ketoreductase determines the organization of the β-carbon processing enzymes of modular polyketide synthases. Structure 14, 737–748 (2006).

    Article  CAS  Google Scholar 

  14. Smith, S. & Tsai, S.C. The type I fatty acid and polyketide synthases: a tale of two megasynthases. Nat. Prod. Rep. 24, 1041–1072 (2007).

    Article  CAS  Google Scholar 

  15. Horton, J.R., Sawada, K., Nishibori, M. & Cheng, X. Structural basis for inhibition of histamine N-methyltransferase by diverse drugs. J. Mol. Biol. 353, 334–344 (2005).

    Article  CAS  Google Scholar 

  16. Tang, Y., Kim, C.Y., Mathews, I.I., Cane, D.E. & Khosla, C. The 2.7-angstrom crystal structure of a 194-kDa homodimeric fragment of the 6-deoxyerythronolide B synthase. Proc. Natl. Acad. Sci. USA 103, 11124–11129 (2006).

    Article  CAS  Google Scholar 

  17. Cheng, Y. et al. Single particle reconstructions of the transferrin-transferrin receptor complex obtained with different specimen preparation techniques. J. Mol. Biol. 355, 1048–1065 (2006).

    Article  CAS  Google Scholar 

  18. Joshi, A.K. & Smith, S. Construction, expression, and characterization of a mutated animal fatty acid synthase deficient in the dehydrase function. J. Biol. Chem. 268, 22508–22513 (1993).

    CAS  PubMed  Google Scholar 

  19. Witkowski, A., Joshi, A.K., Lindqvist, Y. & Smith, S. Conversion of a β-ketoacyl synthase to a malonyl decarboxylase by replacement of the active-site cysteine with glutamine. Biochemistry 38, 11643–11650 (1999).

    Article  CAS  Google Scholar 

  20. Tang, Y., Chen, A.Y., Kim, C.Y., Cane, D.E. & Khosla, C. Structural and mechanistic analysis of protein interactions in module 3 of the 6-deoxyerythronolide B synthase. Chem. Biol. 14, 931–943 (2007).

    Article  CAS  Google Scholar 

  21. Chen, Z.J. et al. Structural enzymological studies of 2-enoyl thioester reductase of the human mitochondrial FAS II pathway: new insights into its substrate recognition properties. J. Mol. Biol. 379, 830–844 (2008).

    Article  CAS  Google Scholar 

  22. Joshi, A.K., Rangan, V.S., Witkowski, A. & Smith, S. Engineering of an active animal fatty acid synthase dimer with only one competent subunit. Chem. Biol. 10, 169–173 (2003).

    Article  CAS  Google Scholar 

  23. Maier, T., Leibundgut, M. & Ban, N. The crystal structure of a mammalian fatty acid synthase. Science 321, 1315–1322 (2008).

    Article  CAS  Google Scholar 

  24. Smith, S. & Abraham, S. Fatty acid synthase from lactating rat mammary gland. Methods Enzymol. 35, 65–74 (1975).

    Article  CAS  Google Scholar 

  25. Tischendorf, G.W., Zeichhardt, H. & Stoffler, G. Determination of the location of proteins L14, L17, L18, L19, L22, L23 on the surface of the 50S ribosomal subunit of Escherichia coli by immune electron microscopy. Mol. Gen. Genet. 134, 187–208 (1974).

    Article  CAS  Google Scholar 

  26. Suloway, C. et al. Automated molecular microscopy: the new Leginon system. J. Struct. Biol. 151, 41–60 (2005).

    Article  CAS  Google Scholar 

  27. Frank, J. et al. SPIDER and WEB: processing and visualization of images in 3D electron microscopy and related fields. J. Struct. Biol. 116, 190–199 (1996).

    Article  CAS  Google Scholar 

  28. Rath, B.K. & Frank, J. Fast automatic particle picking from cryo-electron micrographs using a locally normalized cross-correlation function: a case study. J. Struct. Biol. 145, 84–90 (2004).

    Article  CAS  Google Scholar 

  29. Penczek, P., Radermacher, M. & Frank, J. Three-dimensional reconstruction of single particles embedded in ice. Ultramicroscopy 40, 33–53 (1992).

    Article  CAS  Google Scholar 

  30. Bretaudiere, J.P. & Frank, J. Reconstitution of molecule images analysed by correspondence analysis: a tool for structural interpretation. J. Microsc. 144, 1–14 (1986).

    Article  CAS  Google Scholar 

  31. Radermacher, M., Wagenknecht, T., Verschoor, A. & Frank, J. Three-dimensional reconstruction from a single-exposure, random conical tilt series applied to the 50S ribosomal subunit of Escherichia coli. J. Microsc. 146, 113–136 (1987).

    Article  CAS  Google Scholar 

  32. Keatinge-Clay, A.T. A tylosin ketoreductase reveals how chirality is determined in polyketides. Chem. Biol. 14, 898–908 (2007).

    Article  CAS  Google Scholar 

  33. Pasta, S., Witkowski, A., Joshi, A.K. & Smith, S. Catalytic residues are shared between two pseudosubunits of the dehydratase domain of the animal fatty acid synthase. Chem. Biol. 14, 1377–1385 (2007).

    Article  CAS  Google Scholar 

  34. Soding, J., Biegert, A. & Lupas, A.N. The HHpred interactive server for protein homology detection and structure prediction. Nucleic Acids Res. 33, W244–W248 (2005).

    Article  Google Scholar 

  35. Pettersen, E.F. et al. UCSF Chimera—a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).

    Article  CAS  Google Scholar 

  36. Ploskon, E. et al. A mammalian type I fatty acid synthase acyl carrier protein domain does not sequester acyl chains. J. Biol. Chem. 283, 518–528 (2008).

    Article  CAS  Google Scholar 

  37. Volkmann, N. & Hanein, D. Quantitative fitting of atomic models into observed densities derived by electron microscopy. J. Struct. Biol. 125, 176–184 (1999).

    Article  CAS  Google Scholar 

  38. Koski, M.K., Haapalainen, A.M., Hiltunen, J.K. & Glumoff, T. A two-domain structure of one subunit explains unique features of eukaryotic hydratase 2. J. Biol. Chem. 279, 24666–24672 (2004).

    Article  CAS  Google Scholar 

  39. Oefner, C., Schulz, H., D'Arcy, A. & Dale, G.E. Mapping the active site of Escherichia coli malonyl-CoA-acyl carrier protein transacylase (FabD) by protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 62, 613–618 (2006).

    Article  Google Scholar 

  40. Bunkoczi, G. et al. Mechanism and substrate recognition of human holo ACP synthase. Chem. Biol. 14, 1243–1253 (2007).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank A. Witkowski for helpful discussions. We also acknowledge the National Resource for Automated Molecular Microscopy for assistance with data collection. The work was supported by a research fellowship F32 DK080622 (to E.J.B.) and grant RO1 DK16073 (to S.S.) from the US National Institutes of Health.

Author information

Authors and Affiliations

  1. Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, 92037, California, USA

    Edward J Brignole & Francisco J Asturias

  2. Children's Hospital Oakland Research Institute, 5700 Martin Luther King Jr. Way, Oakland, 94609, California, USA

    Stuart Smith

Authors
  1. Edward J Brignole
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Stuart Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Francisco J Asturias
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

E.J.B. performed all experiments and data analysis; S.S. provided purified FAS; all authors contributed to designing experiments, interpreting results and writing the manuscript.

Corresponding author

Correspondence to Francisco J Asturias.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 (PDF 2633 kb)

Supplementary Video 1

2D analysis reveals a continuum of domain rearrangements within the β-carbon processing portion of FAS. (GIF 2693 kb)

Supplementary Video 2

An animation illustrates how FAS may interchange between the experimentally observed conformations to facilitate contacts between the ACP and active sites of the catalytic domains in a typical reaction cycle. (MOV 9889 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Brignole, E., Smith, S. & Asturias, F. Conformational flexibility of metazoan fatty acid synthase enables catalysis. Nat Struct Mol Biol 16, 190–197 (2009). https://doi.org/10.1038/nsmb.1532

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1532

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing