Native conjugation between proteins and [60]fullerene derivatives using SpyTag as a reactive handle

Herein, we report the facile conjugation between proteins and water-soluble 60]fullerene derivatives (DC₆₀) under native conditions using SpyTag as a reactive handle. Water-soluble 60]fullerene derivatives were first prepared via sequential Bingel-Hirsch reaction and “clicked” with SpyTag to give DC₆₀-SpyTag for native conjugation with proteins by the highly efficient SpyTag-SpyCatcher chemistry. The bioconjugation was confirmed by MALDI-TOF MS spectra and SDS-PAGE analysis. The TEM and UV-vis spectroscopic study further revealed that the DC₆₀ could alter the optical performance and induce aggregation of the target proteins. It thus provides a general and robust method for modifying proteins with C₆₀ derivatives and could potentially be adapted for native conjugation between proteins and other nonbiological motifs as well.