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A chemoselective strategy for late-stage functionalization of complex small molecules with polypeptides and proteins

Nature Chemistry volume 11, pages 78–85 (2019)Cite this article

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Abstract

Conjugates between proteins and small molecules enable access to a vast chemical space that is not achievable with either type of molecule alone; however, the paucity of specific reactions capable of functionalizing proteins and natural products presents a formidable challenge for preparing conjugates. Here we report a strategy for conjugating electron-rich (hetero)arenes to polypeptides and proteins. Our bioconjugation technique exploits the electrophilic reactivity of an oxidized selenocysteine residue in polypeptides and proteins, and the electron-rich character of certain small molecules to provide bioconjugates in excellent yields under mild conditions. This conjugation chemistry enabled the synthesis of peptide–vancomycin conjugates without the prefunctionalization of vancomycin. These conjugates have an enhanced in vitro potency for resistant Gram-positive and Gram-negative pathogens. Additionally, we show that a 6 kDa affibody protein and a 150 kDa immunoglobulin-G antibody could be modified without diminishing bioactivity.

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Fig. 1: The conjugation of oxidized selenocysteine to electron-rich small molecules is a one-pot chemoselective reaction for the covalent attachment of natural products and pharmaceuticals to polypeptides and proteins.
Fig. 2: Selenocysteine-based conjugation of vancomycin to antibacterial peptides enables the discovery of potent Gram-positive and Gram-negative antibacterial agents.
Fig. 3: The conjugation of vancomycin to affibody generates conjugates with retained structures and binding affinities.
Fig. 4: Conjugation of genistain to the antibody leads to conjugates with a retained target binding affinity.
Fig. 5: Selenocysteine-based conjugation of peptides to (hetero)arenes, natural products and pharmaceuticals proceeded in moderate-to-high yields.

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All the data generated or analysed during this study are included in this published article (and in the Supplementary Information). Further details are available from the corresponding authors upon request.

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Acknowledgements

Financial support was provided by NIH Awards GM46059 (S.L.B.), F32GM108294 (D.T.C.), F32GM122204 (L.H.), F30HD093358 (C.M.F.) and GM110535 (B.L.P.) and also by MIT startup funds, a Damon Runyon Cancer Research Foundation Award and a Sontag Distinguished Scientist Award for B.L.P. C.Z. is a recipient of a George Büchi Summer Research Fellowship, a Koch Graduate Fellowship in Cancer Research and a Bristol-Myers Squibb Fellowship in Synthetic Organic Chemistry. A.J.M. is a National Science Foundation Graduate Research Fellow. The authors acknowledge the Biological Instrument Facility of MIT for providing the Octet BioLayer Interferometry System (NIH S10 OD016326) and CD spectrometer (NSF-0070319). We thank Y.-M. Wang and M. Pirnot (MIT) for help in preparing this article. We also acknowledge S. Bano (Merck) for Cu ICP-MS analysis of the purified peptides 13c, 13d, 13e and 13f.

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Author notes

  1. Daniel T. Cohen

    Present address: AbbVie, Inc., North Chicago, IL, USA

  2. These authors contributed equally: Chi Zhang, Colin M. Fadzen.

Authors and Affiliations

  1. Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA

    Daniel T. Cohen, Chi Zhang, Colin M. Fadzen, Alexander J. Mijalis, Stephen L. Buchwald & Bradley L. Pentelute

  2. Department of Chemistry, Yale University, New Haven, CT, USA

    Liana Hie & Scott J. Miller

  3. Visterra Inc., Cambridge, MA, USA

    Kenneth D. Johnson, Zachary Shriver & Obadiah Plante

  4. Koch Institute, Broad Institute of Harvard and MIT, Center for Environmental Health Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA

    Bradley L. Pentelute

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Contributions

D.T.C. and B.L.P. conceived the work and designed the experiments. D.T.C., C.Z. and C.M.F. performed the peptide and protein labelling experiments. D.T.C., C.M.F. and A.J.M. developed the seleno–affibody synthesis. L.H. and S.J.M. performed the NMR characterization to assign regioselectivity for the selenocysteine conjugation. K.D.J., Z.S. and O.P. carried out the cytotoxicity and haemolysis assays on 23v analogues. D.T.C., C.Z., C.M.F. and B.L.P. wrote the manuscript, with input from all other authors.

Corresponding authors

Correspondence to Daniel T. Cohen or Bradley L. Pentelute.

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Competing interests

K.D.J., Z.S. and O.P. are employees of Visterra Inc. D.T.C., C.Z., S.L.B. and B.L.P. are inventors on a patent filed by MIT-TLO to cover this work (US patent application no. 15,187,169 and international application (PCT/US16/38372).

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Cohen, D.T., Zhang, C., Fadzen, C.M. et al. A chemoselective strategy for late-stage functionalization of complex small molecules with polypeptides and proteins. Nature Chem 11, 78–85 (2019). https://doi.org/10.1038/s41557-018-0154-0

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