Homodimeric Protein-Polymer Conjugates via the Tetrazine-trans-Cyclooctene Ligation

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Lorenzo M. M., Decker C. G., Kahveci M. Ü., Paluck S. J., Maynard H. D.

MACROMOLECULES, vol.49, no.1, pp.30-37, 2016 (SCI-Expanded) identifier identifier

  • Publication Type: Article / Article
  • Volume: 49 Issue: 1
  • Publication Date: 2016
  • Doi Number: 10.1021/acs.macromol.5b02323
  • Journal Name: MACROMOLECULES
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus
  • Page Numbers: pp.30-37
  • Istanbul Technical University Affiliated: No


Tetrazine end-functionalized telechelic polymers were synthesized by controlled radical polymerization (CRP) and employed to generate T4 lysozyme homodimers. Mutant T4 lysozyme (V131C), containing a single surface-exposed cysteine, was modified with a protein-reactive trans-cyclooctene (T4L-TCO). Reversible addition-fragmentation chain transfer (RAFT) polymerization yielded poly(N-isopropylacrylamide) (pNIPAAm) with a number-average molecular weight (Mn by 1H NMR) of 2.0 kDa and a dispersity (D by GPC) of 1.05. pNIPAAm was then modified at both ends by postpolymerization with 6-methyltetrazine. For comparison, 2.0 kDa bis-tetrazine poly(ethylene glycol) (PEG) and 2.0 kDa bis-maleimide pNIPAAm were synthesized. Ligation of T4L-TCO to bis-tetrazine pNIPAAm or bis-tetrazine PEG resulted in protein homodimer in 38% yield and 37% yield, respectively, after only 1 h, whereas bis-maleimide pNIPAAm resulted in only 5% yield of dimer after 24 h. This work illustrates the advantage of employing tetrazine ligation over maleimide thiol-ene chemistry for the synthesis of protein homodimer conjugates.