Fig. 3: The pH-sensitive intein performs C-terminal cleavage during EV-biogenesis. | Nature Communications

Fig. 3: The pH-sensitive intein performs C-terminal cleavage during EV-biogenesis.

From: Engineering of extracellular vesicles for efficient intracellular delivery of multimodal therapeutics including genome editors

Fig. 3

a Schematic illustration of the cleavage mechanism of different engineered intein variants. b Protein expression of different engineered mutant intein constructs in whole cell lysates (WCL) and isolated EVs derived from HEK293T cells evaluated by western blot analysis. Lysates from 5 × 105 EV-producing cells and 1×1010 engineered EVs were loaded onto the western blot gel. TSG101, syntenin-1 and β-actin were used as EV markers and Calnexin was used as a cellular organelle marker (endoplasmic reticulum) that should be absent in EV samples. c–e Recombination in reporter cells mediated by EVs derived from engineered cells using mutant inteins (H439Q and N440A) in VEDIC system. Two-way ANOVA (Tukey) multiple comparisons test was used for analysis of (c–e). Experiments were done with 3 biological replicates and data are shown as mean ± SD. a Created in BioRender.com, Zheng, W. (2025) https://BioRender.com/v41e325. Exact statistical analysis was reported in the Source data and Source data are provided as a Source Data file.

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