Efficient Synthesis of Cleavable Cell‐Penetrating Peptide–Cargo Conjugate via Low‐Equivalent of Cell‐Penetrating Peptide Activated by 2,2′‐Dithiodipyridine

Cell-penetrating peptides (CPPs) are favored for protein delivery due to their efficiency, rapidity, and low toxicity. However, conjugation of CPPs to proteins often requires significant amounts of CPPs to ensure yields, which also may result in increased protein dimer formation. Herein, it is reported that the use of low equivalents 2,2'-dithiodipyridine (DPDS)-activated CPP for conjugation cargo allows for high-conversion-rate CPP-cargo conjugates. Using this strategy, high-conversion-rate conjugates of cyclic deca-arginine peptide (cR10) with ubiquitin (Ub) and UbcH7 are obtained using only low equivalents of cR10. Furthermore, three CPPs are successfully conjugated to cargo via DPDS, and their successful cytosolic delivery is confirmed through fluorescence imaging.