Stoichiometric and irreversible cysteine-selective protein modification using carbonylacrylic reagents

  1. Bernardim, B. 36
  2. Cal, P.M.S.D. 35
  3. Matos, M.J. 3
  4. Oliveira, B.L. 3
  5. Martínez-Saéz, N. 3
  6. Albuquerque, I.S. 5
  7. Perkins, E. 1
  8. Corzana, F. 23
  9. Burtoloso, A.C.B. 6
  10. Jiménez-Osés, G. 24
  11. Bernardes, G.J.L. 35
  1. 1 Albumedix Ltd, Castle Court, 59 Castle Boulevard, Nottingham, United Kingdom
  2. 2 Universidad de La Rioja
    info

    Universidad de La Rioja

    Logroño, España

    ROR https://ror.org/0553yr311

  3. 3 University of Cambridge
    info

    University of Cambridge

    Cambridge, Reino Unido

    ROR https://ror.org/013meh722

  4. 4 Universidad de Zaragoza
    info

    Universidad de Zaragoza

    Zaragoza, España

    ROR https://ror.org/012a91z28

  5. 5 Universidade de Lisboa
    info

    Universidade de Lisboa

    Lisboa, Portugal

    ROR https://ror.org/01c27hj86

  6. 6 Universidade de São Paulo
    info

    Universidade de São Paulo

    São Paulo, Brasil

    ROR https://ror.org/036rp1748

Show affiliations +
Journal:
Nature Communications

ISSN: 2041-1723

Year of publication: 2016

Volume: 7

Type: Article

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DOI: 10.1038/NCOMMS13128 SCOPUS: 2-s2.0-84992671846 WoS: WOS:000386218000001 GOOGLE SCHOLAR

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Abstract

Maleimides remain the reagents of choice for the preparation of therapeutic and imaging protein conjugates despite the known instability of the resulting products that undergo thiol-exchange reactions in vivo. Here we present the rational design of carbonylacrylic reagents for chemoselective cysteine bioconjugation. These reagents undergo rapid thiol Michael-addition under biocompatible conditions in stoichiometric amounts. When using carbonylacrylic reagents equipped with PEG or fluorophore moieties, this method enables access to protein and antibody conjugates precisely modified at pre-determined sites. Importantly, the conjugates formed are resistant to degradation in plasma and are biologically functional, as demonstrated by the selective imaging and detection of apoptotic and HER2+ cells, respectively. The straightforward preparation, stoichiometric use and exquisite cysteine selectivity of the carbonylacrylic reagents combined with the stability of the products and the availability of biologically relevant cysteine-tagged proteins make this method suitable for the routine preparation of chemically defined conjugates for in vivo applications. © 2016 The Author(s).