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A mating mechanism to generate diversity for the Darwinian selection of DNA-encoded synthetic molecules

Nature Chemistry volume 14, pages 141–152 (2022)Cite this article

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Abstract

DNA-encoded library technologies enable the screening of synthetic molecules but have thus far not tapped into the power of Darwinian selection with iterative cycles of selection, amplification and diversification. Here we report a simple strategy to rapidly assemble libraries of conformationally constrained peptides that are paired in a combinatorial fashion (suprabodies). We demonstrate that the pairing can be shuffled after each amplification cycle in a process similar to DNA shuffling or mating to regenerate diversity. Using simulations, we show the benefits of this recombination in yielding a more accurate correlation of selection fitness with affinity after multiple rounds of selection, particularly if the starting library is heterogeneous in the concentration of its members. The method was validated with selections against streptavidin and applied to the discovery of PD-L1 binders. We further demonstrate that the binding of self-assembled suprabodies can be recapitulated by smaller (∼7 kDa) synthetic products that maintain the conformational constraint of the peptides.

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Fig. 1: Darwinian evolution by selection/amplification/diversification, Dsuprabody and Psuprabody assemblies, and a general selection-protocol scheme.
Fig. 2: Scheme of PNA–peptide conjugate DNA-templated peptide ligation amenable to suprabody library generation.
Fig. 3: DSuprabody generation.
Fig. 4: Streptavidin selection with the 108-member library.
Fig. 5: PD-L1 selection with a focused library.
Fig. 6: PD-L1 selection with the 108-member library.
Fig. 7: Simulation of selection with or without recombination.

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Data availability

The data supporting the findings of this study are available within this paper and its Supplementary Information. Raw data has been deposited on Zenodo (https://doi.org/10.5281/zenodo.5011513). Source data are provided with this paper.

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Acknowledgements

This work was supported by SNSF grants (169141 and 188406) and the NCCR chemical biology (185898). We thank V. Dutoit (Laboratory of Tumor Immunology, Faculty of Medicine, University of Geneva) for providing the U251 cell line.

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Author notes

  1. These authors contributed equally: B. R. Vummidi, L. Farrera-Soler.

Authors and Affiliations

  1. Department of Organic Chemistry, NCCR Chemical Biology, Faculty of Science, University of Geneva, Geneva, Switzerland

    Balayeshwanth R. Vummidi, Lluc Farrera-Soler, Jean-Pierre Daguer, Millicent Dockerill, Sofia Barluenga & Nicolas Winssinger

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  1. Balayeshwanth R. Vummidi
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  3. Jean-Pierre Daguer
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Contributions

N.W. conceived the technology. B.R.V., L.F.-S., J-.P.D., S.B. and N.W. designed the experiments, analysed data and wrote the manuscript. B.R.V established and performed the selections. L.F.-S. established the chemical ligations. J.-P.D. designed the sequences of the library. B.R.V, L.F.-S. and M.D. synthesized and characterized the hits after selection.

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Correspondence to Nicolas Winssinger.

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The authors declare no competing interests.

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Peer review information Nature Chemistry thanks Ryan Hili, Thomas Kodadek, Alexander Satz and Yixin Zhang for their contribution to the peer review of this work.

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Supplementary information

Supplementary Information

Supplementary information.

Reporting Summary

Supplementary Table 1

Mathematical simulations.

Source data

Source Data Fig. 2

Unprocessed gels for Fig. 2c.

Source Data Fig. 3

Unprocessed gels for Fig. 3a,b.

Source Data Fig. 4

Data for graphs in Fig. 4b–d.

Source Data Fig. 5

Data for graphs in Fig. 5c,e.

Source Data Fig. 6

Data for graphs in Fig. 6d–e.

Source Data Fig. 7

Data for graphs in Fig. 7a–c,e,f.

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Vummidi, B.R., Farrera-Soler, L., Daguer, JP. et al. A mating mechanism to generate diversity for the Darwinian selection of DNA-encoded synthetic molecules. Nat. Chem. 14, 141–152 (2022). https://doi.org/10.1038/s41557-021-00829-5

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