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Post-translational backbone-acyl shift yields natural product-like peptides bearing hydroxyhydrocarbon units

Nature Chemistry volume 14, pages 1413–1420 (2022)Cite this article

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Abstract

Hydroxyhydrocarbon (Hhc) moieties in the backbone of peptidic natural products can exert a substantial influence on the bioactivities of the products, making Hhc units an attractive class of building blocks for de novo peptides. However, despite advances in in vitro genetic code reprogramming, the ribosomal incorporation of Hhc units remains challenging. Here we report a method for in vitro ribosomal synthesis of natural-product-like peptides bearing Hhc units. A series of azide/hydroxy acids were designed as chemical precursors of Hhc units and incorporated into the nascent peptide chain by means of genetic code reprogramming. Post-translational reduction of the azide induced an O-to-N acyl shift to rearrange the peptide backbone. This method allows for one-pot ribosomal synthesis of designer macrocycles bearing various β-, γ- and δ-type Hhc units. We also report the synthesis of a statine-containing peptidomimetic inhibitor of β-secretase 1 as a showcase example.

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Fig. 1: Schematic illustration of the post-translational backbone-acyl shift reaction for in vitro ribosomal synthesis of peptides containing Hhc units.
Fig. 2: Post-translational backbone-acyl shift reaction yielding a γ-peptide linkage on a model peptide (cyPep1-γN3).
Fig. 3: Optimization of the post-translational backbone-acyl shift conditions.
Fig. 4: Scope and limitation of the post-translational backbone-acyl shift to yield various Hhc units.
Fig. 5: In vitro expression of peptides containing statine analogues.
Fig. 6: Application of the post-translational backbone-acyl shift for the in vitro synthesis of the β-secretase 1 inhibitor P10–P4′statV.

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Data availability

Methods and data, including for the synthesis of acyl-donor substrates, characterization of acyl-donor substrates, primer sequences, DNA template assembly schemes, additional LC-MS data for the backbone-acyl shift reactions, MS/MS identification of P10–P4′statV, optimization of flexizyme-mediated acylation conditions and summary of all LC-MS chromatograms discussed in this paper are available in the Supplementary Information. Alternatively, data are available from the corresponding authors upon reasonable request.

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Acknowledgements

We thank H. Murakami for invaluable discussions and helpful suggestions. This work was supported by KAKENHI grants (JP16H06444 to H.S. and Y.G.; JP20H05618 to H.S.; JP17H04762, JP19H01014, JP19K22243 and JP20H02866 to Y.G.; JP19J14230 to T.K.) from the Japan Society for the Promotion of Science and Human Frontier Science Program (to H.S.).

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  1. Department of Chemistry, Graduate School of Science, The University of Tokyo, Bunkyo, Tokyo, Japan

    Tomohiro Kuroda, Yichao Huang, Soichiro Nishio, Yuki Goto & Hiroaki Suga

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  1. Tomohiro Kuroda
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Contributions

Y.G. and H.S. conceived and supervised the study. All authors designed experiments. T.K., Y.H. and Y.G. synthesized acyl-donor substrates. T.K. and S.N. performed expression and modification of AzHyA-containing peptides. All authors analysed the experimental results. T.K., Y.G. and H.S. wrote the manuscript with input from all authors. Y.G. prepared manuscript figures.

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Correspondence to Yuki Goto or Hiroaki Suga.

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Nature Chemistry thanks Jan Vincent Arafiles, Jayanta Chatterjee, Yongchan Kwon and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Kuroda, T., Huang, Y., Nishio, S. et al. Post-translational backbone-acyl shift yields natural product-like peptides bearing hydroxyhydrocarbon units. Nat. Chem. 14, 1413–1420 (2022). https://doi.org/10.1038/s41557-022-01065-1

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