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Structural basis of the Cope rearrangement and cyclization in hapalindole biogenesis

Nature Chemical Biology volume 14, pages 345–351 (2018)Cite this article

Abstract

Hapalindole alkaloids are a structurally diverse class of cyanobacterial natural products defined by their varied polycyclic ring systems and diverse biological activities. These complex metabolites are generated from a common biosynthetic intermediate by the Stig cyclases in three mechanistic steps: a rare Cope rearrangement, 6-exo-trig cyclization, and electrophilic aromatic substitution. Here we report the structure of HpiC1, a Stig cyclase that catalyzes the formation of 12-epi-hapalindole U in vitro. The 1.5-Å structure revealed a dimeric assembly with two calcium ions per monomer and with the active sites located at the distal ends of the protein dimer. Mutational analysis and computational methods uncovered key residues for an acid-catalyzed [3,3]-sigmatropic rearrangement, as well as specific determinants that control the position of terminal electrophilic aromatic substitution, leading to a switch from hapalindole to fischerindole alkaloids.

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Fig. 1: Biogenesis of hapalindole alkaloids.
Fig. 2: HpiC1 structural overview at 1.5 Å.
Fig. 3: Active site of SeMet HpiC1 W73M/K132M.
Fig. 4: In vitro characterization of HpiC1 mutants using 1 as substrate.
Fig. 5: Quantum mechanics analysis.
Fig. 6: Molecular dynamics simulations of the active site.

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Acknowledgements

The authors thank the National Science Foundation under the CCI Center for Selective C-H Functionalization (CHE-1700982), the National Institutes of Health (CA70375 to R.M.W. and D.H.S.), R35 GM118101, and the Hans W. Vahlteich Professorship (to D.H.S.) for financial support. M.G.-B. thanks the Ramón Areces Foundation for a postdoctoral fellowship. J.N.S. acknowledges the support of the National Institute of General Medical Sciences of the National Institutes of Health under Award Number F32GM122218. Computational resources were provided by the UCLA Institute for Digital Research and Education (IDRE) and the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by the NSF (OCI-1053575). The content does not necessarily represent the official views of the National Institutes of Health. Anton 2 computer time was provided by the Pittsburgh Supercomputing Center (PSC) through Grant R01GM116961 from the National Institutes of Health. The Anton 2 machine at PSC was generously made available by D.E. Shaw Research.

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Authors and Affiliations

  1. Life Sciences Institute, University of Michigan, Ann Arbor, MI, USA

    Sean A. Newmister, Shasha Li, Andrew N. Lowell, Fengan Yu, Janet L. Smith & David H. Sherman

  2. Department of Medicinal Chemistry, University of Michigan, Ann Arbor, MI, USA

    Shasha Li & David H. Sherman

  3. Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, CA, USA

    Marc Garcia-Borràs, Jacob N. Sanders, Song Yang & K. N. Houk

  4. Department of Biological Chemistry, University of Michigan, Ann Arbor, MI, USA

    Janet L. Smith

  5. Department of Chemistry, Colorado State University, Fort Collins, CO, USA

    Robert M. Williams

  6. University of Colorado Cancer Center, Aurora, CO, USA

    Robert M. Williams

  7. Department of Chemistry, University of Michigan, Ann Arbor, MI, USA

    David H. Sherman

  8. Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI, USA

    David H. Sherman

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Contributions

S.A.N. conducted protein preparation and crystallography. S.L. cloned the genes and assayed the enzymes. M.G.-B. conducted molecular dynamics simulations. J.N.S. conducted density functional theory calculations. A.N.L. synthesized substrates. F.Y. performed bioinformatics analyses. S.A.N., S.L., M.G.-B., J.N.S., S.Y., J.L.S., R.M.W., K.N.H. and D.H.S. designed research and conducted data analysis and interpretation. S.A.N., S.L., M.G.-B., J.N.S., J.L.S., K.N.H. and D.H.S. wrote the manuscript.

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Correspondence to Robert M. Williams, K. N. Houk or David H. Sherman.

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Newmister, S.A., Li, S., Garcia-Borràs, M. et al. Structural basis of the Cope rearrangement and cyclization in hapalindole biogenesis. Nat Chem Biol 14, 345–351 (2018). https://doi.org/10.1038/s41589-018-0003-x

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