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. 2020 Nov 2;59(45):19951-19955.
doi: 10.1002/anie.202005747. Epub 2020 Sep 2.

Hetero-Diels-Alder Cycloaddition with RAFT Polymers as Bioconjugation Platform

Ana Beloqui  1   2   3   4 , Shivshankar R Mane  1   2 , Marcel Langer  2 , Mathias Glassner  2 , Dennis M Bauer  5 , Ljiljana Fruk  5   6 , Christopher Barner-Kowollik  2   7   8 , Guillaume Delaittre  1   2   9
Affiliations

Hetero-Diels-Alder Cycloaddition with RAFT Polymers as Bioconjugation Platform

Ana Beloqui et al. Angew Chem Int Ed Engl. .

Abstract

We introduce the bioconjugation of polymers synthesized by RAFT polymerization, bearing no specific functional end group, by means of hetero-Diels-Alder cycloaddition through their inherent terminal thiocarbonylthio moiety with a diene-modified model protein. Quantitative conjugation occurs over the course of a few hours, at ambient temperature and neutral pH, and in the absence of any catalyst. Our technology platform affords thermoresponsive bioconjugates, whose aggregation is solely controlled by the polymer chains.

Keywords: Diels-Alder cycloaddition; bioconjugation; end group; polymer; protein.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Scheme 1
Scheme 1
Synthesis of bovin serum albumin polymer conjugates by RAFT‐HDA, as described in the current contribution.
Figure 1
Figure 1
(Top) Synthetic route for oligoethylene glycol‐based polyacrylates by RAFT polymerization in the presence of 2‐cyanoprop‐2‐yl diethoxyphosphoryldithioformate (CPDPDT). (i) mTEGA, AIBN, ethanol, 60 °C. (ii) eDEGA:mOEGA 80/20 mol/mol, AIBN, ethanol, 60 °C. (Bottom) Corresponding SEC traces.
Figure 2
Figure 2
Coomassie‐stained SDS‐PAGE gels of BSA conjugates obtained by RAFT‐HDA with PmTEGA6000 (up) and corresponding electrophoretograms (bottom). (A) Variation of the [PmTEGA6000]:[dBSA] ratio for a fixed reaction time of 12 h: 25 (c), 50 (d), 80 (e), and 100 (f). (B) Variation of the reaction time for a fixed [PmTEGA6000]:[dBSA] ratio of 100: 1 h (g), 2 h (h), 4 h (i), 6 h (j), and 8 h (k). Molecular weight protein ladder (a) and control sample (b, dBSA) are added as references.
Figure 3
Figure 3
Number‐based hydrodynamic diameter distributions for BSA conjugates obtained by RAFT‐HDA with P(eDEGA‐co‐mOEGA) copolymers CoP15000 and CoP18000. dBSA is shown as reference.
Figure 4
Figure 4
(A) Evolution of the turbidity measured at 670 nm with temperature for aqueous solutions of CoP15000 and CoP18000 (15 μM in PBS). (B) Evolution of number‐average hydrodynamic diameters of dBSA‐CoP15000 and dBSA‐CoP18000 conjugates with temperature as measured by DLS. In both cases, measurements were carried out with 1 °C increments and a stabilitization time of 5 min before acquisition.
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