Schulich Faculty of Chemistry, Technion-Israel Institute of Technology, Haifa, 3200008, Israel.
Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Robert-Rössle-Strasse 10, Berlin, 13125, Germany.
Angew Chem Int Ed Engl. 2022 Nov 21;61(47):e202207551. doi: 10.1002/anie.202207551. Epub 2022 Oct 19.
Modifying cyclic cell-penetrating deca-arginine (cR10) peptides with 4-(4-dimethylaminophenylazo)benzoic acid (DABCYL) improves the uptake efficiency of synthetic ubiquitin (Ub) cargoes into living cells. To probe the role of the DABCYL moiety, we performed time-lapse microscopy and fluorescence lifetime imaging microscopy (FLIM) of fluorescent DABCYL-R10 to evaluate the impact on cell entry by the formation of nucleation zones. Furthermore, we performed a structure-uptake relationship study with 13 DABCYL derivatives coupled to CPP to examine their effect on the cell-uptake efficiency when conjugated to mono-Ub through disulfide linkages. Our results show that through structure variations of the DABCYL moiety alone we could reach, at nanomolar concentration, an additional threefold increase in the cytosolic delivery of Ub, which will enable studies on various intracellular processes related to Ub signaling.
用 4-(4-二甲基氨基苯偶氮)苯甲酸（DABCYL）修饰环状细胞穿透性十聚精氨酸（cR10）肽可提高合成泛素（Ub）货物进入活细胞的摄取效率。为了探究 DABCYL 部分的作用，我们通过荧光寿命成像显微镜（FLIM）对荧光 DABCYL-R10 进行了时程显微镜观察，以评估成核区形成对细胞进入的影响。此外，我们还进行了结构摄取关系研究，将 13 种 DABCYL 衍生物与 CPP 偶联，通过二硫键将其与单 Ub 偶联，以检查它们对细胞摄取效率的影响。我们的结果表明，通过单独改变 DABCYL 部分的结构，我们可以在纳摩尔浓度下使 Ub 的胞质内递送增加三倍，这将使与 Ub 信号相关的各种细胞内过程的研究成为可能。
