Development of Endogenous Protein Probes for Characterizing Surface Proteins and Cellular Interactors of Extracellular Vesicles.

ABSTRACT: Extracellular vesicle (EV) surface proteins, derived from producer cells and their surrounding environment, represent a valuable source of biomarkers and participate in a plethora of biological functions, including intercellular communication. However, current methods struggle to distinguish core EV surface proteins from adsorbed corona proteins or map the EV-cell interplay. Here, a genetically encoded proximity labelling probe is presented that displays engineered ascorbate peroxidase, APEX2, on the surface of EVs via fusion to EV-sorting scaffold proteins. This enables the biotinylation of producer-cell-derived surface proteins, corona proteins, and interactors in vitro. After the enrichment by streptavidin bead pulldowns, subpopulation-specific, biotinylated surfaceome and interactome are comprehensively characterized using mass spectrometry-based proteomics. Thus, a genetic tool is introduced for the high-fidelity mapping of the surfaceome and cellular interactome of EVs in vitro. This approach offers a robust framework for dissecting EV biology and has broad applications in biomarker discovery and EV-based therapeutics.

SUBMITTER: Zheng W 

PROVIDER: S-EPMC12948232 | biostudies-literature | 2026 Feb

REPOSITORIES: biostudies-literature