Structural basis of ribosomal peptide macrocyclization in plants

Joel Haywood, Jason W. Schmidberger, Amy M. James, Samuel G. Nonis, Kirill V. Sukhoverkov, Mikael Elias, Charles S. Bond, Joshua S. Mylne

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43 Scopus citations


Constrained, cyclic peptides encoded by plant genes represent a new generation of drug leads. Evolution has repeatedly recruited the Cys-protease asparaginyl endopeptidase (AEP) to perform their head-to-tail ligation. These macrocyclization reactions use the substrates amino terminus instead of water to deacylate, so a peptide bond is formed. How solvent-exposed plant AEPs macrocyclize is poorly understood. Here we present the crystal structure of an active plant AEP from the common sunflower, Helianthus annuus. The active site contained electron density for a tetrahedral intermediate with partial occupancy that predicted a binding mode for peptide macrocyclization. By substituting catalytic residues we could alter the ratio of cyclic to acyclic products. Moreover, we showed AEPs from other species lacking cyclic peptides can perform macrocyclization under favorable pH conditions. This structural characterization of AEP presents a logical framework for engineering superior enzymes that generate macrocyclic peptide drug leads.

Original languageEnglish (US)
Article numbere32955
StatePublished - Jan 31 2018

Bibliographical note

Funding Information:
The authors thank Dan Tawfik for critical insights during project design, Hannes Ludewig for comments on the manuscript and Renier van der Hoorn for the BODIPY probe JOPD1. JSM was supported by an Australian Research Council (ARC) Future Fellowship (FT120100013). SGN and KVS were supported by the Australian Government’s Research Training Program. JH and this work were supported by ARC grant DP160100107 to JSM and Dan Tawfik. This research was undertaken on the MX2 beamline at the Australian Synchrotron, part of the Australian Nuclear Science and Technology Organisation. HaAEP1 is deposited in PDB under accession 6AZT.

Publisher Copyright:
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